Engineered terminal deoxynucleotidyl transferase variants

Abstract

The present invention provides engineered terminal deoxynucleotidyl transferase (TdT) polypeptides useful in template-independent polynucleotide synthesis that are capable of adding, in a template-independent manner, a nucleoside triphosphate-3'-O-removable blocking group (NTP-3'-O-RBG) to the 3'-OH end of a growing oligonucleotide or polynucleotide chain, as well as compositions, methods utilizing these engineered polypeptides, and polynucleotides encoding the engineered terminal deoxynucleotidyl transferases.

Description

[Technical Field]

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 256,353, filed October 15, 2021, and U.S. Provisional Patent Application No. 63 / 329,777, filed April 11, 2022, both of which are incorporated by reference in their entirety for all purposes.

[0002] Technical Field The present invention provides engineered terminal deoxynucleotidyl transferase (TdT) polypeptides useful in template-independent polynucleotide synthesis using nucleoside triphosphate-3'-O-removable blocking groups (NTP-3'-O-RBG), as well as compositions and methods utilizing these engineered polypeptides.

[0003] Reference to a sequence listing, table or computer program An official copy of the Sequence Listing is hereby submitted herewith as an XML file with the filename "CX9-217WO2_ST26.xml", created on October 13, 2022, and 13.3 megabytes in size. The Sequence Listing is a part of the present specification and is incorporated herein by reference in its entirety. [Background technology]

[0004] background Synthetic biology is becoming established in a diverse range of high-value, high-growth markets. From food and agriculture to therapeutics, diagnostics, and vaccines, tools such as gene editing, DNA sequencing, and gene synthesis are being used to construct value-added products with advanced functionality (e.g., cell bioreactors) and desired end products (e.g., drugs, chemicals, etc.). A barrier to the widespread implementation of these technologies is the ability to efficiently synthesize DNA and other polynucleotides. Traditional methods of DNA synthesis use phosphoramidite DNA synthesis chemistry, which is limited to the production of short oligonucleotides of approximately 200 base pairs (Beaucage & Caruthers. (1981). Tetrahedron Lett. 22 (20): 1859). Phosphoramidite DNA synthesis has the additional disadvantage of generating large amounts of toxic waste.

[0005] In the early 1980s, phosphoramidite DNA synthesis was a pivotal development that enabled advances in genomics and enabled new ways to study disease and potential cures. However, over the past 40 years, progress in polynucleotide synthesis methods has been limited and has lagged behind advances in other areas of molecular biology. New methods for synthesizing long, pure strands of natural DNA and other polynucleotides are needed to enable current opportunities in synthetic biology. Template-independent enzymatic synthesis

[0006] Enzymatic synthesis can facilitate the production of long polynucleotides (>200 base pairs) while minimizing toxic waste. Various prokaryotic and eukaryotic DNA and RNA polymerases are known to naturally synthesize polynucleotides of thousands of base pairs or more. Most of these polymerases function during cell division or DNA replication, which involves transcription of DNA to RNA, related to gene or protein expression. Both of these processes involve template-dependent polynucleotide synthesis, in which a polymerase uses a pre-existing template polynucleotide strand to synthesize a complementary polynucleotide strand.

[0007] The potential of template-independent enzymatic polynucleotide synthesis to generate defined sequences has long been recognized. One early report suggested that stepwise addition of specific nucleotide residues was possible using NTPs with blocked 3'-OH groups (Bollum. (1962). JBC, 237, 1945-1949).

[0008] However, several polymerases are known to catalyze template-independent polynucleotide synthesis. These include polymerase lambda, polymerase mu, and terminal deoxynucleotidyl transferase (TdT), all members of the X family of DNA polymerases, many of which are involved in DNA repair processes (Dominguez et al. (2000). EMBO, 19(7), 1731-1742). Among these, TdT is known to generate antigen receptor diversity by indiscriminately adding nucleosides to the 3' end of single-stranded polynucleotides in a template-independent process (Bentolila et al. (1995). EMBO, 14(17), 4221-4229).

[0009] Others have published methods for polynucleotide synthesis using nucleoside 5'-triphosphates with their 3'-OH positions protected by removable blocking moieties, and specifically template-independent polynucleotide polymerases, including terminal deoxynucleotidyl transferase (U.S. Pat. No. 5,763,594). Blocking groups, also known to those skilled in the art as inhibitory groups or terminal groups, can include a variety of groups that block access to the active site, thereby preventing TdT from adding additional NTPs to the nascent polynucleotide chain. These can include charged molecules, large molecules and moieties, or other blocking groups known to those skilled in the art. Suitable removable blocking groups can include carbonitriles, phosphates, carbonates, carbamates, esters, ethers, borates, nitrates, sugars, phosphoramidates, phenylsulfenates, sulfates, and sulfones. Other 3'-OH blocking groups are also known in the art, including 3'-O-amines and methylamines (US Pat. No. 7,544,794), and 3'-O-azides (US Pat. No. 10,407,721).

[0010] Although initially promising, the use of NTPs with a blocked 3'-OH in template-independent synthesis catalyzed by TdT proved difficult in practice because the blocked 3'-OH does not allow the enzyme to recognize the polynucleotide as a substrate for additional synthetic steps.

[0011] Several recent reports have described template-independent synthesis methods using modified NTPs with blocking groups attached to the purine or pyrimidine base, leaving the 3'-OH unmodified and available for additional rounds of synthesis. These base-blocking groups can include cleavable linkers, which allow for removal of the blocking group after each NTP addition step. Cleavable linkers can also be attached to detectable labels (see, inter alia, U.S. Pat. No. 7,057,026). Various cleavable linkers are known to those skilled in the art. These include linkers attached via a reducible disulfide bond, photocleavable, electrophilic or nucleophilic, pH-sensitive, temperature-sensitive, and enzymatically cleavable linkers. One drawback of using cleavable linkers is that some atoms of the linker moiety typically remain attached to the NTP after cleavage, leaving a "scar" that can prevent synthesis of the complementary strand after the initial template-independent synthesis of the primary polynucleotide strand.

[0012] Recently, modified NTPs have been developed that have a base attached to a blocking group via a cleavable linker, leaving the nascent DNA "intact" and ready for the next round of synthesis. In one example, the blocking group and cleavable linker are attached to the base via a disulfide bond. Upon addition of a reducing agent, the blocking group is removed, and the remaining atoms of the linker self-cyclize, leaving the nascent DNA without any linker atoms (U.S. Patent Nos. 8,808,989, 9,695,470, and 10,041,110). Methods for synthesizing polynucleotides using NTPs attached to cleavable blocking groups are known, including using microfluidic devices or inkjet printing technology (U.S. Patent No. 9,279,149). Exonucleases can also be used in methods for synthesizing polynucleotides to shorten or completely degrade polynucleotide chains that did not successfully add NTPs after the polynucleotide elongation step and before removing the blocking group (U.S. Patent No. 9,771,613).

[0013] However, NTP bases labeled with bulky blocking groups attached via cleavable linkers are not optimal for the efficient synthesis of long-chain oligonucleotides. Large labels can negatively affect enzyme kinetics, and damage to the linker can cause unacceptable rates of misincorporation when synthesizing complementary strands. In addition, the larger linkers and the necessary removable steps can increase the cost, time, and inefficiency of the reaction, making these methods economically unfeasible. Recently, several groups have explored modifying the structure or amino acid sequence of TdT or other polymerases to enable template-independent synthesis using 3'-OH blocking groups. Efcavitch et al. described the incorporation of 3'-OH modified dNTPs by TdT in template-independent synthesis using mouse or bacterial TdT with substituted amino acid residues (U.S. Patent No. 10,059,929). Other reports have described engineered bovine and gar (Lepisosteus oculatus) TdTs that showed improved activity relative to wild-type TdT (U.S. Patent No. 10,745,727, PCT / GB2020 / 050247). Similarly, various mutations that improve the activity of Pol X family enzymes have been described (WO2017216472A2). Finally, N-terminal truncation of the BRCT domain of TdT (or alternatively, mutation of the BRCT domain) has also been described to enhance activity in the addition of reversibly blocked NTPs to the 3'-OH of nucleic acids (US20210164008A1). However, despite the recognized value of this technology and intensive research efforts directed toward solving challenges in this field, no feasible method for template-independent enzymatic synthesis of long polynucleotides is currently known or commercially available. Improved engineered TdT enzymes are needed to enable template-independent enzymatic synthesis of long polynucleotides or oligonucleotides of defined sequence using nucleoside triphosphates bearing 3'-O-removable blocking groups. [Prior art documents] [Patent documents]

[0014] [Patent Document 1] U.S. Patent No. 5,763,594 [Patent Document 2] U.S. Patent No. 7,544,794 [Patent Document 3] U.S. Patent No. 10,407,721 [Non-patent literature]

[0015] [Non-Patent Document 1] Beaucage & Caruthers. (1981). Tetrahedron Lett. 22 (20): 1859 [Non-patent document 2] Bollum. (1962). JBC, 237, 1945-1949 [Non-patent document 3] Dominguez et al. (2000). EMBO, 19(7), 1731-1742 [Non-patent document 4] Bentolila et al. (1995). EMBO, 14(17), 4221-4229 Summary of the Invention [Means for solving the problem]

[0016] overview The present invention provides engineered terminal deoxynucleotidyl transferase (TdT) polypeptides useful in template-independent polynucleotide synthesis using nucleoside triphosphate-3'-O-removable blocking groups (NTP-3'-O-RBG), as well as compositions and methods utilizing these engineered polypeptides. The TdT of the present invention is a variant of the predicted splice variant (SEQ ID NO: 2) of the wild-type gene from Empidonax traillii. These engineered TdTs can add nucleoside triphosphates bearing 3'-O-removable blocking groups to the 3'-OH terminus of a growing oligonucleotide or polynucleotide chain in a template-independent manner. After removal of the blocking group, additional rounds of NTP addition can be used to synthesize polynucleotides with defined sequences of bases without using a complementary template strand as a guide for NTP incorporation (template-independent synthesis).

[0017] In some embodiments, the present invention provides a method for the production of nucleic acids comprising SEQ ID NOs: 2, 4, 580, 692, 882, 914, 1034, 1270, 1344, 1346, 1678, 1700, 1750, 1932, 2164, 2666, 2794, 2978, 3074, 3302, 3398, 3488, 3958, 3788, 4124, 4226, 4734, 5052, 5152, 5252, 5296, 5628, 5630 , 5632, and / or 5636, and an engineered TdT polypeptide comprising an amino acid sequence having at least 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% sequence identity to the reference sequences of SEQ ID NOs: 2, 4, 5632, and / or 5636, and containing at least one substitution or one set of substitutions at one or more positions, wherein the positions are selected from SEQ ID NOs: 2, 4, 5632, 5636 ... Numbered with reference to 92, 882, 914, 1034, 1270, 1344, 1346, 1678, 1700, 1750, 1932, 2164, 2666, 2794, 2978, 3074, 3302, 3398, 3488, 3958, 3788, 4124, 4226, 4734, 5052, 5152, 5252, 5296, 5628, 5630, 5632, and / or 5636 The present invention provides engineered TdT polypeptides that have improved thermostability, increased activity at elevated temperatures, increased soluble expression, reduced by-product formation, increased specific activity toward NTP-3'-O-RBG substrates, and / or increased activity toward various oligo acceptor substrates compared to wild-type TdT or other TdTs known to those of skill in the art. These engineered TdT polypeptides with one or more amino acid residue differences or sets of residue differences are described in the detailed description of the invention below.

[0018] In some additional embodiments, the engineered polypeptide comprises an amino acid sequence having at least 60% sequence identity to any of the even-numbered sequences set forth in SEQ ID NOs:6 through 3592 and 3698 through 6766.

[0019] The present invention also provides engineered polynucleotides encoding at least one engineered polypeptide described in the preceding paragraph. In some embodiments, the engineered polynucleotide comprises the odd-numbered sequences set forth in SEQ ID NOs:5 to 3591 and SEQ ID NOs:3697 to 6765.

[0020] The present invention further provides vectors comprising at least one engineered polynucleotide described above. In some embodiments, the vector further comprises at least one regulatory sequence.

[0021] The present invention also provides host cells comprising the vectors provided herein. In some embodiments, the host cells produce at least one engineered polypeptide provided herein.

[0022] The present invention further provides methods for producing an engineered TdT polypeptide, comprising culturing a host cell provided herein under conditions such that the engineered polynucleotide is expressed and the engineered polypeptide is produced. In some embodiments, the method further comprises recovering the engineered polypeptide. [Brief explanation of the drawings]

[0023] [Figure 1] Figure 1 depicts the relationships of the full-length TdT variants and the enzymes based on sequence conservation as a phylogenetic tree.

[0024] [Figure 2] Figure 2 depicts the relationships of the truncated TdT variants and enzymes based on sequence conservation as a phylogenetic tree.

[0025] [Figure 3-1] Figures 3A, 3B, 3C, and 3D represent alignments of TdT variants, highlighting conserved residues. [Figure 3-2] Figures 3A, 3B, 3C, and 3D represent alignments of TdT variants, highlighting conserved residues. [Figure 3-3] Figures 3A, 3B, 3C, and 3D represent alignments of TdT variants, highlighting conserved residues. [Figure 3-4] Figures 3A, 3B, 3C, and 3D represent alignments of TdT variants, highlighting conserved residues.

[0026] [Figure 4] FIG. 4 depicts an evaluation of TAATC binding and detection, as described in Example 6 below. DETAILED DESCRIPTION OF THE INVENTION

[0027] Description of the Invention Unless otherwise defined, all technical and scientific terms used herein generally have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Generally, the nomenclature used herein and the laboratory procedures of cell culture, molecular genetics, microbiology, organic chemistry, analytical chemistry, and nucleic acid chemistry described below are those well known and commonly used in the art. Such techniques are well known and described in numerous texts and references well known to those skilled in the art. Standard techniques, or modifications thereof, are used for chemical synthesis and chemical analysis. All patents, patent applications, documents, and publications mentioned herein, both supra and infra, are hereby expressly incorporated herein by reference.

[0028] Although any suitable methods and materials similar or equivalent to those described herein can be used in the practice of the present invention, some methods and materials are described herein. It should be understood that the present invention is not limited to the specific methodology, protocols, and reagents described, as they may vary depending on the circumstances used by those skilled in the art. Thus, the terms defined immediately below are more fully described with reference to the present invention as a whole.

[0029] It should be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the present invention. The section headings used herein are for organizational purposes only and should not be construed as limiting the subject matter described. Numerical ranges are inclusive of the numbers defining that range. Accordingly, every numerical range disclosed herein is intended to include every narrower numerical range that falls within a broader numerical range, as if such narrower numerical ranges were all expressly written herein. Every maximum (or minimum) numerical limitation disclosed herein is also intended to include every lower (or higher) numerical limitation, as if such lower (or higher) numerical limitations were expressly written herein.

[0030] As used herein and in the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a polypeptide" includes two or more polypeptides. Similarly, the terms "comprise," "comprises," "comprising," "include," "includes," and "including" are interchangeable and are not intended to be limiting.

[0031] When the description of various embodiments uses the term "comprising," it should be understood that one of ordinary skill in the art would understand that in some specific instances, the embodiment could instead be described using the words "consisting essentially of" or "consisting of." When the description of various embodiments uses the terms "optionally" and "optionally," it is further understood that the subsequently described event or circumstance may or may not occur, and that the description includes examples in which the event or circumstance occurs and examples in which it does not occur. It is understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not limiting of the present disclosure. The section headings used herein are for organizational purposes only and should not be construed as limiting the subject matter described. Abbreviation:

[0032] The abbreviations used for the genetically encoded amino acids are conventional and are as follows: [Table 112]

[0033] When three-letter abbreviations are used, the amino acid may be in either the L- or D-configuration about the alpha carbon (Cα) unless specifically preceded by "L" or "D" or unless otherwise clear from the context in which the abbreviation is used. For example, "Ala" refers to alanine without specifying the configuration about the alpha carbon, while "D-Ala" and "L-Ala" refer to D-alanine and L-alanine, respectively.

[0034] When single-letter abbreviations are used, an uppercase letter refers to an amino acid in the L-configuration about the α-carbon, and a lowercase letter refers to an amino acid in the D-configuration about the α-carbon. For example, "A" refers to L-alanine and "a" refers to D-alanine. When a polypeptide sequence is represented as a series of single-letter or three-letter abbreviations (or mixtures thereof), the sequence is represented in the amino (N) to carboxy (C) direction, in accordance with common convention.

[0035] The abbreviations used for genetically coded nucleosides are conventional and are as follows: adenosine (A); guanosine (G); cytidine (C); thymidine (T); and uridine (U). These abbreviations are also used interchangeably for nucleosides and nucleotides (nucleosides having one or more phosphate groups). Unless specifically specified, the abbreviated nucleoside or nucleotide may be either a ribonucleoside (or ribonucleotide) or a 2'-deoxyribonucleoside (or 2'-deoxyribonucleotide). Nucleosides or nucleotides may also be modified at the 3' position. Nucleosides or nucleotides may be individually or collectively identified as either a ribonucleoside (or ribonucleotide) or a 2'-deoxyribonucleoside (or 2'-deoxyribonucleotide). When a nucleic acid sequence is represented as a series of single-letter abbreviations, the sequence is represented, in accordance with common convention, in the 5' to 3' direction and the phosphate is not indicated.

[0036] definition With respect to the present invention, technical and scientific terms used in the description herein have the meanings commonly understood by those skilled in the art, unless otherwise defined. Accordingly, the following terms are intended to have the following meanings:

[0037] The "EC" number refers to the Enzyme Nomenclature of the International Union of Biochemistry and Molecular Biology (NC-IUBMB) Committee on Nomenclature. The IUBMB biochemical classification is a numerical classification system for enzymes based on the chemical reaction they catalyze.

[0038] "ATCC" refers to the American Type Culture Collection, whose biorepository collection includes genes and strains.

[0039] "NCBI" refers to the National Center for Biotechnology Information and the sequence databases it hosts.

[0040] "Protein," "polypeptide," and "peptide" are used interchangeably herein to refer to a polymer of at least two amino acids covalently linked by an amide bond, regardless of length or post-translational modification (e.g., glycosylation, phosphorylation, lipidation, myristoylation, ubiquitination, etc.). Included within this definition are D- and L-amino acids and mixtures of D- and L-amino acids, as well as polymers comprising D- and L-amino acids and mixtures of D- and L-amino acids.

[0041] "Amino acids" are referred to herein by either their commonly known three letter symbols or by the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Similarly, nucleotides may be referred to by their commonly accepted single-letter symbols.

[0042] As used herein, "polynucleotide," "oligonucleotide," and "nucleic acid" are used interchangeably herein and refer to two or more nucleosides or nucleotides covalently linked together. A polynucleotide may be composed entirely of ribonucleotides (i.e., RNA), entirely of 2'-deoxyribonucleotides (i.e., DNA), entirely of other synthetic nucleotides, or a mixture of synthetic ribonucleotides and / or 2'-deoxyribonucleotides. A polynucleotide may also contain nucleotides with substitutions, including 2'-substitutions (e.g., 2'-fluoro or 2'-O-methyl). Nucleosides are typically linked together via standard phosphodiester bonds, although a polynucleotide may contain one or more non-standard linkages (including, but not limited to, phosphothiolated linkages). A polynucleotide may be single-stranded or double-stranded, or may contain both single-stranded and double-stranded regions. Additionally, polynucleotides are typically composed of naturally occurring coding nucleobases (i.e., adenine, guanine, uracil, thymine, and cytosine), but may also contain one or more modified and / or synthetic nucleobases, such as, for example, inosine, xanthine, hypoxanthine, etc. In some embodiments, such modified or synthetic nucleobases are nucleobases that encode an amino acid sequence. Modified or synthetic nucleobases may include any known, hypothetical, or future-discovered modification or structure recognized by those of skill in the art as a modified or synthetic nucleobase. Similarly, the terms "polynucleotide," "oligonucleotide," and "nucleic acid" are intended to include any currently known or future-discovered modified or synthetic structure recognized by those of skill in the art as being or having the functionality of a "polynucleotide," "oligonucleotide," or "nucleic acid." An example of a modified or synthetic structure having the functionality of a "polynucleotide," "oligonucleotide," or "nucleic acid" is PNA, or peptide nucleic acid.

[0043] As used herein, "oligo acceptor substrate," "acceptor substrate," "growing oligo acceptor substrate strand," and "growing polynucleotide strand" are used interchangeably herein and refer to any oligo or nucleotide strand, or similar moiety with an exposed 3'-OH, that can be recognized by wild-type TdT or polymerases, or engineered TdT or polymerases of the present disclosure, as a substrate for nucleoside addition or synthesis. In some embodiments, the acceptor substrate may be single-stranded. In still other embodiments, the acceptor substrate may be double-stranded or partially double-stranded. In some embodiments, the acceptor substrate may comprise a nucleotide strand consisting of 1-10 NTPs, 5-20 NTPs, 15-50 NTPs, 30-100 NTPs, or more than 100 NTPs. In some embodiments, the acceptor substrate may not be a nucleotide strand, but may comprise a chemical moiety containing a free -OH that can be recognized as a substrate by wild-type or engineered TdT. Exemplary oligo acceptor substrates are provided in the Examples.

[0044] As used herein, "nucleoside triphosphate-3'-O-removable blocking group" and "nucleotide triphosphate-3'-O-removable blocking group" and "NTP-3'-O-RBG" are used interchangeably herein and refer to a ribonucleoside triphosphate or deoxyribonucleoside triphosphate, or a synthetic or nucleoside triphosphate, composed of an alternative or modified sugar having a removable blocking group attached at the 3'OH of the sugar moiety.

[0045] As used herein, "oligo acceptor product" and "oligo acceptor extension product" are used interchangeably herein and refer to the product of an NTP-3'-O-RBG substrate and an oligo acceptor substrate, where TdT or a related polymerase has catalyzed the extension or addition of NTP-3'-O-RBG to the oligo acceptor substrate.

[0046] As used herein, "removable blocking group," "blocking group," "terminating group," and "inhibitory group," as well as related variations of these terms, are used interchangeably herein and refer to a chemical group that prevents the addition of a second NTP-3'-O-RBG substrate to the 3' end of a growing oligo acceptor substrate chain prior to removal from the first round of addition of the removable blocking group. In some embodiments, the NTP-3'-O-RBG substrate may contain a removable blocking group selected from the group consisting of NTP-3'-O-NH2, or NTP-3'-O-PO3. In some embodiments, the NTP-3'-O-RBG may have a natural purine or pyrimidine base, such as adenine, guanine, cytosine, thymine, or uridine. In some embodiments, the NTP-3'-O-RBG may have an unnatural base analog, such as inosine, xanthine, hypoxanthine, or another base analog known in the art.

[0047] As used herein, " template-independent synthesis " refers to the synthesis of oligonucleotide or polynucleotide without using a template strand as a guide for the synthesis of complementary oligonucleotide or polynucleotide strand.Therefore, template-independent synthesis refers to an iterative process, whereby successful NTP is added to growing oligonucleotide or nucleotide strand, or to acceptor substrate.Template-independent synthesis can be sequence-defined, or can be random, as in the case of using wild-type TdT in generating diversity of antigen receptors.The process for template-independent synthesis is further described herein.

[0048] "Coding sequence" refers to a portion of a nucleic acid (eg, a gene) that codes for the amino acid sequence of a protein.

[0049] "Naturally occurring" or "wild-type" refers to a form found in nature. For example, a naturally occurring or wild-type polypeptide or polynucleotide sequence is a sequence present in an organism that can be isolated from a natural source and has not been intentionally modified by human manipulation.

[0050] As used herein, "recombinant," "engineered," and "non-naturally occurring," when used in reference to a cell, nucleic acid, or polypeptide, refer to material that has been altered in a way that would not otherwise occur in nature, or material that corresponds to the natural or native form of the material. In some embodiments, the cell, nucleic acid, or polypeptide is identical to a naturally occurring cell, nucleic acid, or polypeptide, but is produced or obtained from synthetic materials and / or by manipulation using recombinant techniques. Non-limiting examples include recombinant cells that express genes not found within the native (non-recombinant) form of the cell, or that express native genes that are otherwise expressed at different levels, among others.

[0051] "Percentage of sequence identity" and "percentage of homology" are used interchangeably herein to refer to a comparison between polynucleotides or polypeptides and are determined by comparing two optimally aligned sequences over a comparison window, where the portion of the polynucleotide or polypeptide sequence in the comparison window may contain additions or deletions (i.e., gaps) compared to the reference sequence due to optimal alignment of the two sequences. The percentage may be calculated by determining the number of positions where the same nucleic acid base or amino acid residue is present in both sequences to obtain the number of matched positions, dividing the number of matched positions by the total number of positions in the comparison window, and multiplying the result by 100 to obtain the percentage of sequence identity. Alternatively, the percentage may be calculated by determining the number of positions where either the same nucleic acid base or amino acid residue is present in both sequences, or by aligning the nucleic acid base or amino acid residue with gaps to obtain the number of matched positions, dividing the number of matched positions by the total number of positions in the comparison window, and multiplying the result by 100 to obtain the percentage of sequence identity. Those skilled in the art are well aware that there are many established algorithms available for aligning two sequences.The optimal alignment of the sequences for comparison can be carried out, for example, by Smith and Waterman's local homology algorithm (Smith and Waterman, Adv. Appl. Math., 2:482

[1981] ), Needleman and Wunsch's homology alignment algorithm (Needleman and Wunsch, J. Mol. Biol., 48:443

[1970] ), Pearson and Lipman's similarity search method (Pearson and Lipman, Proc. Natl. Acad. Sci. USA 85:2444

[1988] ), computerized implementations of these algorithms (for example, GAP, BESTFIT, FASTA and TFASTA in the GCG Wisconsin software package), or visual inspection, as known in the art.Examples of algorithms suitable for determining percent sequence identity and percent sequence similarity include, but are not limited to, the BLAST and BLAST 2.0 algorithms described by Altschul et al. (see Altschul et al., J. Mol. Biol., 215: 403-410

[1990] ; and Altschul et al., Nucl. Acids Res., 3389-3402

[1977] , respectively). Software for performing BLAST analyses is publicly available through the website of the National Center for Biotechnology Information. This algorithm involves identifying high-scoring sequence pairs (HSPs) by first identifying short word lengths W in the query sequence that, when aligned with words of the same length in a database sequence, either match or meet some positive threshold score T. T is referred to as the neighborhood word score threshold (Altschul et al., supra). These initial neighborhood word hits act as seeds for initiating searches to find longer HSPs containing them. The word hits are then extended in both directions along each sequence as far as possible to increase the cumulative alignment score. For nucleotide sequences, the cumulative score is calculated using the parameters M (reward score for a pair of matching residues; always >0) and N (penalty score for mismatching residues; always <0). For amino acid sequences, a scoring matrix is ​​used to calculate the cumulative score. Extension of the word hits in each direction is stopped when: the cumulative alignment score falls by an amount X from its maximum achieved value; the cumulative score falls to zero or below due to the accumulation of one or more negative-scoring residue alignments; or the end of either sequence is reached. The BLAST algorithm parameters W, T, and X determine the sensitivity and speed of the alignment. The BLASTN program (for nucleotide sequences) uses as defaults a word length (W) of 11, an expectation (E) of 10, M=5, N=-4, and a comparison of both strands.For amino acid sequences, the BLASTP program uses as defaults a word length (W) of 3, an expectation (E) of 10, and the BLOSUM62 scoring matrix (see Henikoff and Henikoff, Proc. Natl. Acad. Sci. USA 89:10915

[1989] ). Exemplary sequence alignments and determination of percent sequence identity can utilize the BESTFIT or GAP programs in the GCG Wisconsin software package (Accelrys, Madison WI) using the default parameters provided.

[0052] A "reference sequence" refers to a defined sequence used as a basis for sequence and / or activity comparison. A reference sequence may be a subset of a larger sequence, e.g., a segment of a full-length gene or polypeptide sequence. Generally, a reference sequence is at least 20 nucleotides or amino acid residues in length, at least 25 residues in length, at least 50 residues in length, or the full-length nucleic acid or polypeptide. Because two polynucleotides or polypeptides may each contain (1) sequences that are similar between the two sequences (i.e., a portion of the complete sequence) and (2) additional sequences that differ between the two sequences, sequence comparison between two (or more) polynucleotides or polypeptides is typically performed by comparing the sequences of the two polynucleotides or polypeptides over a "comparison window" to identify and compare local regions of sequence similarity. In some embodiments, a "reference sequence" can be based on a primary amino acid sequence, where the reference sequence may have one or more changes in the primary sequence. For example, a "reference sequence based on SEQ ID NO: 4 with a valine at the residue corresponding to X14," or X14V, refers to a reference sequence in which the residue corresponding to X14 in SEQ ID NO: 4, which is a tyrosine, has been changed to a valine.

[0053] A "comparison window" refers to a conceptual segment of at least about 20 contiguous nucleotide positions or amino acid residues, where a sequence may be compared to a reference sequence of at least 20 contiguous nucleotides or amino acids, and the portion of the sequence in the comparison window may contain 20 percent or fewer additions or deletions (i.e., gaps) compared to the reference sequence (no additions or deletions) for optimal alignment of the two sequences. The comparison window may be longer than 20 contiguous residues, including windows of 30, 40, 50, 100, or longer, as appropriate.

[0054] As used herein, "substantial identity" refers to a polynucleotide or polypeptide sequence that shares at least 80 percent sequence identity, at least 85 percent sequence identity, at least 89-95 percent sequence identity, or more usually at least 99 percent sequence identity with respect to a reference sequence over a comparison window of at least 20 residue positions, often over a window of at least 30-50 residues, where the percentage of sequence identity is calculated by comparing the reference sequence to a sequence that contains a total of 20 percent or fewer deletions or additions of the reference sequence over the comparison window. In certain embodiments, as applied to polypeptides, the term "substantial identity" means that two polypeptide sequences share at least 80 percent sequence identity, preferably at least 89 percent sequence identity, at least 95 percent or more sequence identity (e.g., 99 percent sequence identity), when optimally aligned, such as by the programs GAP or BESTFIT, using default gap weights. In some embodiments, non-identical residue positions in the compared sequences differ by conservative amino acid substitutions.

[0055] "Corresponding," "with reference to," and "compared to," when used in the context of the numbering of a given amino acid or polynucleotide sequence, refer to the numbering of residues in a specified reference sequence when the given amino acid or polynucleotide sequence is compared to the reference sequence. In other words, the residue number or residue position of a given polymer is specified with respect to the reference sequence rather than by the actual numerical position of the residue within the given amino acid or polynucleotide sequence. For example, a given amino acid sequence, such as the amino acid sequence of an engineered TdT, can be aligned with a reference sequence by introducing gaps to optimize residue matching between the two sequences. In these cases, although gaps exist, the numbering of residues in a given amino acid or polynucleotide sequence is performed with respect to the reference sequence to which it is aligned.

[0056] An "amino acid difference" or "residue difference" or "substitution" refers to a change in an amino acid residue at a position in a polypeptide sequence compared to the amino acid residue at the corresponding position in a reference sequence. The position of an amino acid difference is generally referred to herein as "Xn," where n refers to the corresponding position in the reference sequence at which the residue difference is based. For example, a "residue difference at position X25 compared to SEQ ID NO:2" refers to a change in the amino acid residue at the polypeptide position corresponding to position 25 of SEQ ID NO:2. Thus, if a reference polypeptide of SEQ ID NO:2 has a valine at position 25, then a "residue difference at position X25 compared to SEQ ID NO:2" refers to an amino acid substitution of any residue other than serine at the polypeptide position corresponding to position 25 of SEQ ID NO:2. In most examples herein, a specific amino acid residue difference at a position will be designated as "XnY," where "Xn" identifies the corresponding position as described above, and "Y" is the single-letter identifier of the amino acid found in the engineered polypeptide (i.e., the residue that differs from the reference polypeptide). In some embodiments, more than one amino acid may appear at a particular residue position (i.e., alternative amino acids may be listed in the form XnY / Z, where Y and Z represent alternative amino acid residues).In some cases (e.g., Tables 7.2, 8.2, 9.2, 10.2, 11.2, 12.2, 13.2, 14.2, 15.2, 16.2, 17.2, 18.2, 19.2, 20.2, 21.2, 22.2, 23.2, 24.2, 25.2, 26.2, 27.2, 28.2, 29.2, 30.2, 31.2, 32.2, 33.2, 34.2, 35.2, 36 .2, 37.2, 38.2, 39.2, 40.2, 41.2, 42.2, 43.2, 44.2, 45.2, 46.2, 47.2, 48.2, 49.2, 50.2, 51.2, 52.2, 53.2, 54.2, 55.2, 56.2, 57.2, 58.2, 59.2, 60.2, 61.2, 62.2, 63.2, 64.2, 65.2, 66.2, 74. 2, 75.2, 76.2, 77.2, 78.2, 79.2, 80.2, 81.2, 82.2, 83.2, 84.2, 85.2, 86.2, 87.2, 88.2, 89.2, 90.2, 91.2, 92.2, 93.2, 94.2, 95.2, 96.2, 97.2, 98.2, 99.2, 100.2, 101.2, 102.2, 103.2, 104.2 , 105.2, 106.2, 107.2, and 108.2), the present invention also provides specific amino acid differences, represented by the conventional designation "AnB," where A is the single-letter identifier of the residue in the reference sequence, "n" is the number of the residue position in the reference sequence, and B is the single-letter identifier of the residue substitution in the engineered polypeptide sequence. Furthermore, in some examples, the polypeptides of the invention can include one or more amino acid residue differences compared to the reference sequence, represented by a list of specified positions where changes were made compared to the reference sequence. In some additional embodiments, the present invention provides engineered polypeptide sequences that include both conservative and non-conservative amino acid substitutions.

[0057] As used herein, "conservative amino acid substitution" refers to the replacement of a residue with a different residue having a similar side chain, and thus typically includes the replacement of an amino acid in a polypeptide with an amino acid within the same or similar defined amino acid class.By way of example and not limitation, an amino acid with an aliphatic side chain is replaced with another aliphatic amino acid (e.g., alanine, valine, leucine, and isoleucine); an amino acid with a hydroxyl side chain is replaced with another amino acid with a hydroxyl side chain (e.g., serine and threonine); an amino acid with an aromatic side chain is replaced with another amino acid with an aromatic side chain (e.g., phenylalanine, tyrosine, tryptophan, and histidine); an amino acid with a basic side chain is replaced with another amino acid with a basic side chain (e.g., lysine and arginine); an amino acid with an acidic side chain is replaced with another amino acid with an acidic side chain (e.g., aspartic acid or glutamic acid); and / or a hydrophobic or hydrophilic amino acid is replaced with another hydrophobic or hydrophilic amino acid, respectively.Exemplary conservative substitutions are provided in Table 1 below. [Table 1]

[0058] A "non-conservative substitution" refers to the replacement of an amino acid in a polypeptide with an amino acid having significantly different side chain properties. Non-conservative substitutions may use amino acids between, rather than within, a defined group and affect (a) the structure of the peptide backbone in the area of ​​the substitution (e.g., proline for glycine), (b) the charge or hydrophobicity, or (c) the bulk of the side chain. By way of example and not limitation, exemplary non-conservative substitutions may be an acidic amino acid substituted with a basic or aliphatic amino acid, an aromatic amino acid substituted with a small amino acid, and a hydrophilic amino acid substituted with a hydrophobic amino acid.

[0059] "Deletion" refers to a modification to a polypeptide by removing one or more amino acids from a reference polypeptide. Deletions can include the removal of one or more amino acids, two or more amino acids, five or more amino acids, ten or more amino acids, fifteen or more amino acids, or twenty or more amino acids, up to 10% of the total number of amino acids comprising the reference enzyme, or up to 20% of the total number of amino acids, while retaining the enzymatic activity and / or improved properties of the engineered TdT enzyme. Deletions can be directed to internal and / or terminal portions of the polypeptide. In various embodiments, deletions can include a continuous segment or can be discontinuous.

[0060] "Insertion" refers to a modification to a polypeptide by the addition of one or more amino acids from a reference polypeptide. In some embodiments, the improved engineered TdT enzyme comprises an insertion of one or more amino acids relative to a naturally occurring polypeptide, as well as an insertion of one or more amino acids relative to other improved TdT polypeptides. The insertion can be in the internal portion of the polypeptide, or at the carboxy- or amino-terminus. As used herein, insertion includes fusion proteins known in the art. The insertion can be a continuous segment of amino acids, or can be separated by one or more amino acids in a naturally occurring polypeptide.

[0061] "Fragment," as used herein, refers to a polypeptide having an amino- and / or carboxy-terminal deletion, but where the remaining amino acid sequence is identical to the corresponding positions in the sequence. Fragments can be at least 14 amino acids long, at least 20 amino acids long, at least 50 amino acids long, or longer, and can be up to 70%, 80%, 90%, 95%, 98%, and 99% of a full-length TdT polypeptide, e.g., the polypeptide of SEQ ID NO: 4, or the TdT provided in the even-numbered sequences of SEQ ID NOs: 6-3592 and 3698-6766.

[0062] An "isolated polypeptide" refers to a polypeptide that has been substantially separated from other contaminants (e.g., proteins, lipids, and polynucleotides) that naturally accompany it. This term encompasses polypeptides that have been removed from their naturally occurring environment or expression system (e.g., host cell or in vitro synthesis) or that have been purified. Engineered TdT enzymes may be present within cells, in cell culture media, or prepared in various forms, such as a lysate or isolated preparation. Thus, in some embodiments, an engineered TdT enzyme may be an isolated polypeptide.

[0063] "Substantially pure polypeptide" refers to a composition in which the polypeptide species is the predominant species present (i.e., it is more abundant than any other individual macromolecular species in the composition, on a molar or weight basis). A composition is generally substantially purified when the desired species constitutes at least about 50 percent of the macromolecular species present, by molar or weight percent. Generally, a substantially pure TdT composition will comprise about 60% or more, about 70% or more, about 80% or more, about 90% or more, about 95% or more, and about 98% or more of all macromolecular species present in the composition, by molar or weight percent. In some embodiments, the desired species is purified to essential homogeneity (i.e., contaminant species cannot be detected in the composition by conventional detection methods), where the composition consists essentially of a single macromolecular species. Solvent species, small molecules (<500 Daltons), and elemental ion species are not considered macromolecular species. In some embodiments, an isolated engineered TdT polypeptide is a substantially pure polypeptide composition.

[0064] As used herein, "improved enzymatic property" refers to at least one improved property of an enzyme. In some embodiments, the present invention provides engineered TdT polypeptides that exhibit an improvement in any enzymatic property compared to a reference TdT polypeptide, and / or a wild-type TdT polypeptide, and / or another engineered TdT polypeptide. For the engineered TdT polypeptides described herein, the comparison is generally made to the wild-type enzyme from which the TdT is derived, although in some embodiments, the reference enzyme can be another improved engineered TdT. Thus, the level of "improvement" can be determined and compared among various TdT polypeptides, including wild-type as well as engineered TdT. Improved properties include, but are not limited to, properties such as enzyme activity (which may be expressed in terms of percent conversion of substrate), thermostability, solvent stability, pH activity profile, cofactor requirements, resistance to inhibitors (e.g., substrate or product inhibition), activity at elevated temperatures, increased soluble expression, reduced by-product formation, increased specific activity toward the NTP-3'-O-RBG substrate, and / or increased activity (including enantioselectivity) toward various oligo acceptor substrates.

[0065] "Increased enzyme activity" refers to an improved property of a TdT polypeptide, which can be expressed as an increase in specific activity (e.g., product produced / time / weight protein) or an increase in percent conversion of substrate to product (e.g., percent conversion of a starting amount of substrate to product over a specified period of time using a specified amount of TdT) compared to a reference TdT enzyme. Exemplary methods for determining enzyme activity are provided in the Examples. m , V max or k catAny property related to enzyme activity, including the classical enzyme properties of the TdT polypeptide, can be affected, and the change can result in increased enzyme activity. Improvements in enzyme activity can range from about 1.2-fold that of the corresponding wild-type enzyme to 2-fold, 5-fold, 10-fold, 20-fold, 25-fold, 50-fold, or even greater enzyme activity than naturally occurring or another engineered TdT from which the TdT polypeptide is derived. TdT activity can be measured by any one of standard assays, such as by monitoring changes in the properties of the substrate, cofactor, or product. In some embodiments, the amount of product produced can be measured by liquid chromatography-mass spectrometry (LC-MS), HPLC, or other methods known in the art. Comparisons of enzyme activity are performed using defined properties of the enzyme, defined assays under set conditions, and one or more defined substrates, as further described in detail herein. Generally, when lysates are compared, not only are the number of cells and amount of protein assayed determined, but the same expression system and the same host cells are used to minimize variation in the amount of enzyme produced by the host cells and present in the lysates.

[0066] "Conversion" refers to the enzymatic conversion of a substrate(s) to the corresponding product(s). "Percent conversion" refers to the percent of a substrate that is converted to a product under specific conditions within a given period of time. Thus, the "enzymatic activity" or "activity" of a TdT polypeptide can be expressed as the "percent conversion" of substrate to product.

[0067] "Thermostable" refers to a TdT polypeptide that maintains similar activity (e.g., 60% to 80% greater) after exposure to elevated temperatures (e.g., 40 to 80°C) for a period of time (e.g., 0.5 to 24 hours) compared to a wild-type enzyme exposed to the same elevated temperature.

[0068] "Solvent-stable" refers to a TdT polypeptide that maintains similar activity (e.g., 60% to 80% greater) after exposure to different concentrations (e.g., 5 to 99%) of a solvent (e.g., ethanol, isopropyl alcohol, dimethyl sulfoxide (DMSO), tetrahydrofuran, 2-methyltetrahydrofuran, acetone, toluene, butyl acetate, methyl tert-butyl ether, etc.) for a period of time (e.g., 0.5 to 24 hours) compared to a wild-type enzyme exposed to the same concentration of the same solvent.

[0069] "Heat and solvent stable" refers to a TdT polypeptide that is both heat and solvent stable.

[0070] The term "stringent hybridization conditions" as used herein refers to conditions under which nucleic acid hybrids are stable. As known to those skilled in the art, hybrid stability is determined by the melting temperature (T m ) Generally, hybrid stability is a function of ionic strength, temperature, G / C content, and the presence of chaotropic agents. mValues ​​can be calculated using known methods for predicting melting temperatures (e.g., Baldino et al., Meth. Enzymol., 168:761-777

[1989] ; Bolton et al., Proc. Natl. Acad. Sci. USA 48:1390

[1962] ; Bresslauer et al., Proc. Natl. Acad. Sci. USA 83:8893-8897

[1986] ; Freier et al., Proc. Natl. Acad. Sci. USA 83:9373-9377

[1986] ; Kierzek et al., Biochem., 25:7840-7846

[1986] ; Rychlik et al., 1990, Nucl. Acids Res., 18:6409-6412

[1990] ). (See, e.g., Nucl. Acids Res., 19:698

[1991] ; Sambrook et al., supra; Suggs et al., 1981, in Developmental Biology Using Purified Genes, Brown et al. [eds.], pp. 683-693, Academic Press, Cambridge, MA

[1981] ; and Wetmur, Crit. Rev. Biochem. Mol. Biol. 26:227-259

[1991] .) In some embodiments, the polynucleotide encodes a polypeptide disclosed herein and hybridizes under defined conditions, such as moderately stringent or highly stringent conditions, to the complement of a sequence encoding an engineered TdT enzyme of the invention.

[0071] "Hybridization stringency" refers to hybridization conditions, such as washing conditions, in nucleic acid hybridization. Generally, hybridization reactions are performed under conditions of lower stringency, followed by washing at various but higher stringencies. The term "moderately stringent hybridization" refers to conditions that allow target DNA to bind to complementary nucleic acids with greater than about 90% identity to the target polynucleotide, and with about 60% identity, preferably about 75% identity, or about 85% identity to the target DNA. Exemplary medium-stringent conditions are equivalent to hybridization in 50% formamide, 5x Denhardt's solution, 5x SSPE, 0.2% SDS at 42°C, followed by washing in 0.2x SSPE, 0.2% SDS at 42°C. "High stringency hybridization" generally refers to hybridization at a thermal melting temperature T determined under solution conditions for a defined polynucleotide sequence. m To about 10°C or lower. In some embodiments, high stringency conditions refer to conditions that allow hybridization of only nucleic acid sequences that form stable hybrids in 0.018M NaCl at 65°C (i.e., if a hybrid is not stable in 0.018M NaCl at 65°C, it is not stable under high stringency conditions as contemplated herein). High stringency conditions can be provided, for example, by hybridization in 50% formamide, 5x Denhardt's solution, 5x SSPE, 0.2% SDS, conditions equivalent to 42°C, followed by washing in 0.1x SSPE and 0.1% SDS at 65°C. Another high stringency condition is hybridization under conditions equivalent to hybridization in 5×SSC containing 0.1% (w / v) SDS at 65° C., and washing in 0.1×SSC containing 0.1% SDS at 65° C. Other high stringency and moderate stringency hybridization conditions are described in the references cited above.

[0072] A "heterologous" polynucleotide refers to any polynucleotide that is introduced into a host cell by laboratory techniques, and includes polynucleotides that are removed from a host cell, subjected to laboratory manipulation, and then reintroduced into the host cell.

[0073] "Codon optimization" refers to changing the codons of a polynucleotide encoding a protein to those preferentially used in a particular organism so that the encoded protein can be efficiently expressed in the organism of interest. Although the genetic code is degenerate in that most amino acids are represented by several codons called "synonymous" or "synonymous" codons, it is well known that the codon usage frequency by a particular organism is not random but is biased toward certain codon triplets. This codon usage bias may be higher in association with a given gene, genes of common function or ancestral origin, highly expressed proteins relative to low copy number proteins, and aggregated protein-coding regions of an organism's genome. In some embodiments, a polynucleotide encoding a TdT enzyme may be codon-optimized for optimal production from the host organism selected for expression.

[0074] As used herein, "preferred, optimal, high codon usage bias codons" refer interchangeably to codons that are used in protein-coding regions at a higher frequency than other codons that encode the same amino acid. Preferred codons can be determined with respect to a single gene, a set of genes of common function or origin, codon usage in highly expressed genes, codon frequency in aggregate protein-coding regions across organisms, codon frequency in aggregate protein-coding regions of related organisms, or a combination thereof. Codons whose frequency increases with the level of gene expression are typically optimal codons for expression. Various methods are known for determining codon frequency (e.g., codon usage frequency, relatively synonymous codon usage frequency) and codon preference in a particular organism, including, for example, cluster analysis or correspondence analysis, and multivariate analysis using the effective number of codons used in a gene (e.g., GCG CodonPreference, Genetics Computer Group Wisconsin Package; CodonW, Peden, University of Nottingham; McInerney, Bioinform., 14:372-73

[1998] ; Stenico et al. (See, e.g., Wada et al., Nucl. Acids Res., 20:2111-2118

[1992] ; Nakamura et al., Nucl. Acids Res., 28:292

[2000] ; Duret et al., supra; Henaut and Danchin, in Escherichia coli and Salmonella, Neidhardt et al. (eds.), ASM Press, Washington DC, pp. 2047-2066

[1996] ). The data source for obtaining codon usage can rely on any available nucleotide sequence capable of coding for a protein.These datasets include nucleic acid sequences that are actually known to encode expressed proteins (e.g., complete protein-coding sequences - CDS), expressed sequence tags (ESTS), or predicted coding regions of genomic sequences (see, e.g., Mount, Bioinformatics: Sequence and Genome Analysis, Chapter 8, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY

[2001] ; Uberbacher, Meth. Enzymol., 266:259-281

[1996] ; and Tiwari et al., Comput. Appl. Biosci., 13:263-270

[1997] ).

[0075] "Control sequences" is defined herein to include all components necessary or advantageous for the expression of polynucleotides and / or polypeptides of the present invention. Each control sequence may be native or foreign to the nucleic acid sequence encoding the polypeptide. Such control sequences include, but are not limited to, a leader, polyadenylation sequence, propeptide sequence, promoter, signal peptide sequence, and transcription terminator. At a minimum, control sequences include a promoter, and transcriptional and translational stop signals. Control sequences may be provided with linkers for the purpose of introducing specific restriction sites to facilitate ligation of the control sequences with the coding region of the nucleic acid sequence encoding the polypeptide.

[0076] "Operably linked" is defined herein as an arrangement in which control sequences are positioned appropriately (i.e., in a functional relationship) relative to a polynucleotide of interest so that the control sequences direct or regulate the expression of the polynucleotide and / or polypeptide of interest.

[0077] A "promoter sequence" refers to a nucleic acid sequence recognized by a host cell for expression of a polynucleotide of interest, such as a coding sequence. The promoter sequence contains transcriptional control sequences that mediate expression of the polynucleotide of interest. The promoter may be any nucleic acid sequence that exhibits transcriptional activity in the host cell of choice, including mutant, truncated, and hybrid promoters, and may be obtained from genes encoding extracellular or intracellular polypeptides that are either homologous or heterologous to the host cell.

[0078] "Suitable reaction conditions" refer to conditions in a biocatalytic reaction solution (e.g., ranges of enzyme loading, substrate loading, cofactor loading, temperature, pH, buffer, cosolvent, etc.) under which a TdT polypeptide of the invention can convert one or more substrate compounds into product compounds (e.g., addition of NTP-3'-O-RBG to an oligo acceptor substrate). Exemplary "suitable reaction conditions" are provided herein and illustrated by the Examples.

[0079] "Composition" refers to a mixture or combination of one or more substances, where each substance or component of the composition retains its individual properties. As used herein, a biocatalytic composition refers to a combination of one or more substances useful for biocatalysis.

[0080] As used herein, "addition amount," such as in "compound addition amount" or "enzyme addition amount" or "cofactor addition amount," refers to the concentration or amount of a component in the reaction mixture at the start of the reaction.

[0081] "Substrate," in the context of a biocatalyst-mediated process, refers to the compound or molecule that is acted upon by the biocatalyst. For example, the TdT biocatalyst used in the synthetic processes disclosed herein acts on an NTP-3'-O-RBG substrate and an oligo acceptor substrate.

[0082] "Product" in the context of a biocatalyst-mediated process refers to a compound or molecule that results from the action of the biocatalyst. For example, exemplary products for the TdT biocatalyst used in the processes disclosed herein are the oligo acceptor extension products depicted in Schemes 1 and 2.

[0083] "Alkyl" refers to a saturated hydrocarbon group of 1 to 18 carbon atoms, inclusive, either straight or branched, more preferably 1 to 8 carbon atoms, inclusive, and most preferably 1 to 6 carbon atoms, inclusive. Alkyl groups having a specific number of carbon atoms are indicated in parentheses (e.g., (C1-C6) alkyl refers to alkyl of 1 to 6 carbon atoms).

[0084] "Alkenyl" refers to a hydrocarbon group of 2 to 12 carbon atoms, inclusive, either straight or branched, containing at least one double bond, but optionally containing two or more double bonds.

[0085] "Alkynyl" refers to a hydrocarbon group of 2 to 12 carbon atoms, inclusive, either straight or branched, containing at least one triple bond, and optionally containing two or more triple bonds, and additionally optionally containing one or more double bond moieties.

[0086] "Heteroalkyl," "heteroalkenyl," and "heteroalkynyl" refer to alkyl, alkenyl, and alkynyl, respectively, as defined herein, in which one or more carbon atoms are each independently replaced with the same or different heteroatom or heteroatom group. Heteroatoms and / or heteroatom groups that can replace carbon atoms include, but are not limited to, -O-, -S-, -SO-, -NR □ -, -PH-, -S(O)-, -S(O)2-, -S(O)NR □ -, -S(O)NR □ -, etc. (including combinations thereof), wherein each R □□ is independently selected from hydrogen, alkyl, cycloalkyl, heterocycloalkyl, aryl and heteroaryl.

[0087] "Amino" refers to the group -NH2. Substituted amino refers to the group -NHR □ , N.R. □ R □ , and N.R. □ R □ R □ In the formula, each R □ are independently selected from substituted or unsubstituted alkyl, cycloalkyl, cycloheteroalkyl, alkoxy, aryl, heteroaryl, heteroarylalkyl, acyl, alkoxycarbonyl, sulfanyl, sulfinyl, sulfonyl, etc. Exemplary amino groups include, but are limited to, dimethylamino, diethylamino, trimethylammonium, triethylammonium, methylsulfonylamino, furanyl-oxy-sulfamino, and the like.

[0088] "Aminoalkyl" refers to an alkyl group in which one or more hydrogen atoms are replaced with one or more amino groups, including substituted amino groups.

[0089] "Aminocarbonyl" refers to -C(O)NH2. Substituted aminocarbonyl is -C(O)NR □ R □ In the formula, the amino group NR □ R □ is as defined herein.

[0090] "Oxy" refers to the divalent group -O- which may have a variety of substituents to form different oxy groups including ethers and esters.

[0091] "Alkoxy" or "alkyloxy" are used interchangeably herein and refer to a group -OR □ In the formula, R □ is an alkyl group, including optionally substituted alkyl groups.

[0092] "Carboxy" refers to --COOH.

[0093] "Carbonyl" refers to -C(O)-, which may have a variety of substituents to form different carbonyl groups including acids, acid halides, aldehydes, amides, esters, and ketones.

[0094] "Carboxyalkyl" refers to an alkyl in which one or more hydrogen atoms are replaced by one or more carboxy groups.

[0095] "Aminocarbonylalkyl" refers to an alkyl substituted with an aminocarbonyl group, as defined herein.

[0096] "Halogen" and "halo" refer to fluoro, chloro, bromo and iodo.

[0097] "Haloalkyl" refers to an alkyl group in which one or more hydrogen atoms have been replaced with halogen. Thus, the term "haloalkyl" is meant to include monohaloalkyl, dihaloalkyl, trihaloalkyl, etc., up to perhaloalkyl. For example, the expression "(C1-C2)haloalkyl" includes 1-fluoromethyl, difluoromethyl, trifluoromethyl, 1-fluoroethyl, 1,1-difluoroethyl, 1,2-difluoroethyl, 1,1,1 trifluoroethyl, perfluoroethyl, etc.

[0098] "Hydroxy" refers to --OH.

[0099] "Hydroxyalkyl" refers to an alkyl group in which one or more hydrogen atoms are replaced by one or more hydroxy groups.

[0100] A "thiol" or "sulfanyl" refers to -SH. A substituted thiol or sulfanyl is -SR □ In the formula, R □is an alkyl, aryl, or other suitable substituent.

[0101] "Sulfonyl" refers to -SO2-. Substituted sulfonyl is -SO2-R □ In the formula, R □ is an alkyl, aryl, or other suitable substituent.

[0102] "Alkylsulfonyl" is -SO2-R □ In the formula, R □ is alkyl, which may be optionally substituted. Typical alkylsulfonyl groups include, but are not limited to, methylsulfonyl, ethylsulfonyl, n-propylsulfonyl, and the like.

[0103] "Phosphate," as used herein, refers to a functional group composed of an orthophosphate ion (a phosphorus atom covalently linked to four oxygen atoms), which is typically found with one or more hydrogen atoms or organic groups.

[0104] "Phosphorylated," as used herein, refers to the addition or presence of one or more phosphoryl groups (a phosphorus atom covalently linked to three oxygen atoms).

[0105] "Optionally substituted," as used herein with respect to the aforementioned chemical groups, means that a position of that chemical group occupied by a hydrogen may be replaced with another atom exemplified by, but not limited to, carbon, oxygen, nitrogen, or sulfur (unless otherwise specified), or with, but not limited to, hydroxy, oxo, nitro, methoxy, ethoxy, alkoxy, substituted alkoxy, trifluoromethoxy, haloalkoxy, fluoro, chloro, bromo, iodo, halo, methyl, ethyl, propyl, butyl, alkyl, alkenyl, alkynyl, substituted alkyl, trifluoromethyl, haloalkyl, hydroxyalkyl, alkoxyalkyl, thio, alkylthio, acyl, carboxy, alkoxycarbonyl, carboxamido, substituted carboxamido, alkylsulfonyl, alkylsulfinyl, alkylsulfonylamino, sulfonamido, substituted sulfonamido, cyano, This means that the heteroatoms can be substituted with chemical groups exemplified by amino, substituted amino, alkylamino, dialkylamino, aminoalkyl, acylamino, amidino, amidoximo, hydroxamoyl, phenyl, aryl, substituted aryl, aryloxy, arylalkyl, arylalkenyl, arylalkynyl, pyridyl, imidazolyl, heteroaryl, substituted heteroaryl, heteroaryloxy, heteroarylalkyl, heteroarylalkenyl, heteroarylalkynyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloalkyl, cycloalkenyl, cycloalkylalkyl, substituted cycloalkyl, cycloalkyloxy, pyrrolidinyl, piperidinyl, morpholino, heterocycle, (heterocycle)oxy, and (heterocycle)alkyl, where preferred heteroatoms are oxygen, nitrogen, and sulfur.Furthermore, if open valences exist on these substituent chemical groups, they can be further substituted with alkyl groups, cycloalkyl groups, aryl groups, heteroaryl groups, and / or heterocyclic groups; if these open valences exist on carbon, they can be further substituted with halogens and with substituents attached to oxygen, nitrogen, or sulfur; and if multiple such open valences exist, these groups can be linked to form rings, either by direct bond formation or by forming a bond to a new heteroatom, preferably oxygen, nitrogen, or sulfur. It is further contemplated that the replacement of hydrogen with a substituent can be made provided that the replacement does not introduce unacceptable instability into the molecules of the present invention and is otherwise chemically reasonable. Those skilled in the art will understand that for any chemical group described as optionally substituted, only sterically practical and / or synthetically feasible chemical groups are meant to be included. As used herein, "optionally substituted" refers to all subsequent modifiers in a term or series of chemical groups. For example, in the term "optionally substituted arylalkyl," the "alkyl" and "aryl" portions of the molecule can be substituted or unsubstituted, and for the series "optionally substituted alkyl, cycloalkyl, aryl, and heteroaryl," the alkyl, cycloalkyl, aryl, and heteroaryl groups can, independently of the other, be substituted or unsubstituted.

[0106] "Reaction," as used herein, refers to a process in which one or more substances or compounds or substrates are converted into one or more different substances or compounds or processes. Template-independent synthesis by engineered TdT

[0107] New methods for efficiently synthesizing long, pure strands of DNA and other polynucleotides are needed to overcome the limitations of existing phosphoramidite chemical synthesis methods in order to enable a range of emerging and existing synthetic biology applications.

[0108] The present invention provides novel terminal deoxynucleotidyl transferases with improved activity in the template-independent synthesis of polynucleotides using 5'-nucleoside triphosphates ("NTPs") modified with a 3'-O-removable blocking group (NTP-3'-O-RBG). The TdTs of the present disclosure have improved thermostability, activity at elevated temperatures, increased soluble expression, reduced by-product formation, increased affinity for NTP-3'-O-RBG substrates, increased affinity for oligoacceptor substrates, increased activity or specific activity for NTP-3'-O-RBG substrates, and / or increased activity or specific activity for various oligoacceptor substrates, compared to wild-type TdT or other TdTs known to those skilled in the art. The engineered polypeptides of the present disclosure are variants of SEQ ID NO: 2, a predicted splice variant encoded by the genome of the species Empidonax traillii. These engineered TdTs are capable of template-independent synthesis of oligonucleotides and polynucleotides.

[0109] Template-independent synthesis of defined polynucleotide sequences using engineered TdT is a multi-step process. In one embodiment, an oligo acceptor substrate bearing a 3'-OH allows for the addition of a defined NTP-3'-O-RBG substrate by the engineered TdT, as depicted in Scheme 1 below. [ka] and B = adenine, guanine, cytidine, thymine, uracil, pseudouridine, 1-methylpseudouridine R1 = H; 6-fluorescein or other fluorescein dye; alkynyl, thiol, amino, biotin or other linker R2 = benzyl, O-nitrobenzyl, benzoyl, acetyl, cyanoethyl, NH2, NO2, PO3 2 - R3=H, OH, OMe, F, Br, Cl, I, Me R4=H, OH, OMe, F, Br, Cl, I, Me X=O, S

[0110] After addition of NTP-3'-O-RBG to the 3'-OH of an oligo acceptor substrate or growing polynucleotide chain, TdT is blocked from further addition by 3'-O-RBG. RBG is then removed, exposing the 3'-OH and allowing another round of addition. After each round of addition, the NTP-3'-O-RBG substrate from the previous round is removed and a new NTP-3'-O-RBG substrate is added, sequentially and efficiently creating defined polynucleotide sequences by addition at the 3'-OH terminus of the primer-free polynucleotide or oligo acceptor substrate.

[0111] A variety of oligo acceptor substrates and NTP-3'-O-RBG substrates can be used in this process, as would be expected by one skilled in the art. One example reaction is detailed in Scheme 2 below. Scheme 2 depicts the TdT-catalyzed reaction of [N]ATC and 3'-phos-dCTP, as described in Example 15, although other examples of suitable oligo acceptor substrate and NTP-3'-O-RBG pairs are described in other examples. These examples are non-limiting. [ka]

[0112] Occasionally, unwanted synthesis products are generated by TdT during the addition step, including incorporation of NTPs that have lost their blocking groups, addition of more than one NTP, or removal or "backchewing" of TdT in the growing polynucleotide chain.

[0113] In some embodiments, one or more additional quality control steps, such as the addition of an exonuclease, are used before removing the blocking groups and initiating a new round of synthesis. In some embodiments, a phosphatase is used to degrade inorganic phosphate and drive the reversible TdT reaction toward synthesis.

[0114] As further described herein, the engineered TdT polypeptides of the present disclosure exhibit one or more improved properties in the template-independent polynucleotide synthesis process depicted in Scheme 1.

[0115] In some embodiments, the present invention provides engineered TdT polypeptides comprising an amino acid sequence having at least 60% sequence identity to the amino acid reference sequence of SEQ ID NO: 4 and further comprising one or more amino acid residue differences compared to the reference amino acid sequence, wherein the engineered TdT polypeptide has improved thermostability, increased activity at elevated temperatures, increased soluble expression, reduced by-product formation, increased specific activity toward NTP-3'-O-RBG substrates, and / or increased activity toward various oligo acceptor substrates compared to wild-type TdT or other TdTs known to those of skill in the art.

[0116] In particular, the engineered TdT polypeptides of the present disclosure have been engineered for the efficient synthesis of long polynucleotides using NTP-3'-O-RBG in the processes described above.

[0117] A variety of suitable reaction conditions will be known to those skilled in the art, as detailed below and in the Examples. Engineered terminal deoxynucleotidyl transferase polypeptides

[0118] The present invention provides engineered terminal deoxynucleotidyl transferase (TDT) polypeptides useful in template-independent polynucleotide synthesis using nucleoside triphosphate-3'-O-removable blocking groups, as well as compositions and methods utilizing these engineered polypeptides.

[0119] The present invention provides TdT polypeptides, polynucleotides encoding the polypeptides, methods of preparing the polypeptides, and methods for using the polypeptides. Where a description refers to a polypeptide, it should be understood that this may also describe a polynucleotide encoding the polypeptide.

[0120] Suitable reaction conditions under which the improved properties of the engineered polypeptides described above are desired can be determined with respect to conditions including polypeptide concentration or amount, substrate, co-substrate, buffer, solvent, pH, temperature and reaction time, and / or conditions using polypeptides immobilized on a solid support, as further described below and in the Examples.

[0121] In some embodiments, exemplary engineered TdTs are listed in Table 7.2, 8.2, 9.2, 10.2, 11.2, 12.2, 13.2, 14.2, 15.2, 16.2, 17.2, 18.2, 19.2, 20.2, 21.2, 22.2, 23.2, 24.2, 25.2, 26.2, 27.2, 28.2, 29.2, 30.2, 31.2, 32.2, 33.2, 34.2, 35.2, 36.2, 37.2, 38.2, 39.2, 40.2, 41.2, 42.2, 43.2, 44.2, 45.2, 46.2, 47.2, 48.2, 49.2, 50.2, 51.2, 52.2, 53.2, 54.2, 55.2, 56.2, 57.2, 58.2, 59.2, 60.2, 61.2, 62.2, 63.2, 64.2, 65.2, 66.2, 67.2, 68.2, 69.2, 70.2, 71.2, 72.2, 73.2, 74.2, 75.2, 76.2, 77.2, 78.2, 79.2, 80.2, 81.2, 82.2, 83.2, 84.2, 85.2, 86.2, 87.2, 88.2, 89.2, .2, 31.2, 32.2, 33.2, 34.2, 35.2, 36.2, 37.2, 38.2, 39.2, 40.2, 41.2, 42.2, 43.2, 44.2, 45.2, 46.2, 47.2, 48.2, 49.2, 50.2, 51.2, 52.2, 53.2, 54.2, 55.2, 56.2, 57.2, 58.2, 59.2, 60.2, 61.2, 62.2, 63.2, 64.2, 65.2, 66.2, 67.2, 68.2, 69.2, 70.2, 71.2, 72.2, 73.2, 74.2, 75.2, 76.2, 77.2, 78.2, 79.2, 80.2, 81.2, 82.2, 83.2, 84.2, 85.2, 86.2, 87.2, 88.2, 89.2, 90.2, 91.2, 92.2, 93.2, 94.2, 95.2, 96.2, 97.2, 98.2, 99.2, 100.2, 101.2, 102.2, 103.2, 104.2, 105.2, 106.2, 107.2, 108.2, 109.2, 110.2, 111.2, 112.2, 1 8.2, 59.2, 60.2, 61.2, 62.2, 63.2, 64.2, 65.2, 66.2, 74.2, 75.2, 76.2, 77.2, 78.2, 79.2, 80.2, 81.2, 82.2, 83.2, 84.2, 85.2, 86.2, 87.2, 88.2, 89.2, 90.2, 91.2, 92.2, and an amino acid sequence having one or more residue differences compared to SEQ ID NO:4 at residue positions set forth as 93.2, 94.2, 95.2, 96.2, 97.2, 98.2, 99.2, 100.2, 101.2, 102.2, 103.2, 104.2, 105.2, 106.2, 107.2, and 108.2.

[0122] Structural and functional information for exemplary engineered polypeptides of the invention is based on the conversion of oligo acceptor substrates and NTP-3'-O-RBG or dideoxyNTPs, the results of which are listed in Tables 7.2, 8.2, 9.2, 10.2, 11.2, 12.2, 13.2, 14.2, 15.2, 16.2, 17.2, 18.2, 19.2, 20.2, 21.2, 22.2, 23.2, 24.2, 25.2, 26.2, 27.2, 28.2, 29.2, 30.2, 31.2, 32.2, 33.2, 34.2, 35.2, 36.2, 37.2, 38.2, 39.2, 40.2, 41.2, 42.2, 43.2, 44.2, 45.2, 46.2, 47.2, 48.2, 49.2, 50.2, 51.2, 52.2, 53.2, 54.2, 55.2, 56.2, 57.2, 58.2, 59.2, 60.2, 61.2, 62.2, 63.2, 64.2, 65.2, 66.2, 67.2, 68.2, 69.2, 70.2, 71.2, 72.2, 73.2, 74.2, 75.2, 76.2, 77.2, 78.2, 79.2, 80.2, 81.2, 82.2, 83. .2, 46.2, 47.2, 48.2, 49.2, 50.2, 51.2, 52.2, 53.2, 54.2, 55.2, 56.2, 57.2, 58.2, 59.2, 60.2, 61.2, 62.2, 63.2, 64.2, 65.2, 66.2, 74.2, 75.2, 76.2, 77.2, 78.2, 79.2, 80.2, 81.2, 82.2, 83.2, 84.2, 85.2, 86.2, 87.2, 88.2, 89.2, 90.2, 91.2, 92.2, 93.2, 94.2, 95.2, 96.2, 97.2, 98.2, 99.2, 100.2, 101.2, 102.2, 103.2, 104.2, 105.2, 106.2, 107.2, 108.2, 109.2, 110.2, 111.2, 112.2, 113.2, 114.2, 115.2, 116.2, 117.2, 118.2, 119.2, 120.2, 121.2, 122.2, 123.2, 124.2, 125.2, 126.2, 127.2, 128.2, 129.2, 130.2, 131.2, The amino acid sequences of the amino acid sequences provided by the even-numbered SEQ ID NOs in these tables are shown in Tables 2.2, 83.2, 84.2, 85.2, 86.2, 87.2, 88.2, 89.2, 90.2, 91.2, 92.2, 93.2, 94.2, 95.2, 96.2, 97.2, 98.2, 99.2, 100.2, 101.2, 102.2, 103.2, 104.2, 105.2, 106.2, 107.2, and 108.2. Odd-numbered sequence identifiers (i.e., SEQ ID NOs) in these tables refer to nucleotide sequences that encode the amino acid sequences provided by the even-numbered SEQ ID NOs in these tables. Exemplary sequences are provided in the electronic sequence listing submitted accompanying this application, which is hereby incorporated by reference herein. Amino acid residue differences are based on comparison to the reference sequence of SEQ ID NOs: 2, 4, 580, 692, 882, 914, 1034, 1270, 1344, 1346, 1678, 1700, 1750, 1932, 2164, 2666, 2794, 2978, 3074, 3302, 3398, 3488, 3958, 3788, 4124, 4226, 4734, 5052, 5152, 5252, 5296, 5628, 5630, 5632, and / or 5636, as indicated.

[0123] Terminal deoxynucleotidyl transferases, members of the Pol X family, have been identified in many species. Diverse Pol X family members are known to share certain residues that are conserved across family members. TdT also has a high level of conservation across species for residues thought to be involved in divalent metal ion binding, ternary complex formation, and dNTP and DNA ligand binding (Dominguez et al. (2000). EMBO, 19(7), 1731-1742). Additionally, TdT is known to have splice variants that lack the BRCT domain and are N-terminal truncations.

[0124] In the present disclosure, TdT from various species, including both truncated and full-length enzymes, was screened for soluble expression, as described below in Example 72. Figure 1 represents the full-length TdT variants and the relationship of the enzymes based on sequence conservation as a phylogenetic tree. Figure 2 represents the truncated TdT variants and the relationship of the enzymes based on sequence conservation as a phylogenetic tree. Figures 3A, 3B, 3C, and 3D represent alignments of TdT from various species screened for soluble expression in the present disclosure, highlighting conserved residues between these species.

[0125] Based on this screen, a predicted splice variant of wild-type TdT from Empidonax traillii (SEQ ID NO: 2) was selected based on soluble expression. The TdT polypeptide of the present disclosure is an engineered variant of SEQ ID NO: 2 and SEQ ID NO: 4, which is an N-terminal 6-histidine tagged version of WT TdT truncated at amino acid position 131 (SEQ ID NO: 4).

[0126] The polypeptides of the present disclosure have residue differences that result in the improved properties necessary to develop efficient TdT enzymes capable of template-independent synthesis of long polynucleotides. Various residue differences at both conserved and non-conserved positions have been discovered to be associated with improved properties of the enzyme, including improved thermostability, increased activity at elevated temperatures, increased soluble expression, reduced by-product formation, increased specific activity toward NTP-3'-O-RBG substrates, and / or increased activity toward various oligoacceptor substrates, compared to wild-type TdT or other TdTs known to those skilled in the art.

[0127] The activity of each engineered TdT relative to the reference polypeptide of SEQ ID NO: 2, 4, 580, 692, 882, 914, 1034, 1270, 1344, 1346, 1678, 1700, 1750, 1932, 2164, 2666, 2794, 2978, 3074, 3302, 3398, 3488, 3958, 3788, 4124, 4226, 4734, 5052, 5152, 5252, 5296, 5628, 5630, 5632, and / or 5636 was determined as substrate conversion as described in the Examples herein. In some embodiments, shake-flask purified enzyme (SFP) was used as a secondary screen to evaluate the properties of the engineered TdT, the results of which are provided in the Examples.

[0128] In some embodiments, specific enzymatic properties are associated with residue differences at residue positions set forth herein compared to SEQ ID NOs: 2, 4, 580, 692, 882, 914, 1034, 1270, 1344, 1346, 1678, 1700, 1750, 1932, 2164, 2666, 2794, 2978, 3074, 3302, 3398, 3488, 3958, 3788, 4124, 4226, 4734, 5052, 5152, 5252, 5296, 5628, 5630, 5632, and / or 5636. In some embodiments, residue differences that affect polypeptide expression can be used to increase expression of the engineered TdT.

[0129] Given the guidance provided herein, exemplary manipulations involving even numbered sequences of SEQ ID NOs: 2, 4, 580, 692, 882, 914, 1034, 1270, 1344, 1346, 1678, 1700, 1750, 1932, 2164, 2666, 2794, 2978, 3074, 3302, 3398, 3488, 3958, 3788, 4124, 4226, 4734, 5052, 5152, 5252, 5296, 5628, 5630, 5632, and / or 5636 are contemplated. Any of the polypeptides listed in Tables 7.2, 8.2, 9.2, 10.2, 11.2, 12.2, 13.2, 14.2, 15.2, 16.2, 17.2, 18.2, 19.2, 20.2, 21.2, 22.2, 23.2, 24.2, 25.2, 26.2, 27.2, 28.2, 29.2, 30.2, 31.2, 32.2, 33.2, 34.2, 35.2, 36.2, 37.2, 38.2, 39.2, 40.2, 41.2, 42.2, 43.2, 44.2, 45.2, 46.2, 47.2, 48.2, 49.2, 50.2, 51.2, 52.2, 53.2, 54.2, 55.2, 56.2, 57.2, 58.2, 59.2, 60.2, 61.2, 62.2, 63.2, 64.2, 65.2, 66.2, 74.2, 75.2, 76.2, 77.2, 78.2, 79.2, 80.2, 81.2, 82.2, 83.2, 84.2, 85.2, 86.2, 87.2, 88.2, 89.2, 90.2, 91.2, 92.2, 93. It is further contemplated that the amino acid sequences of 100.2, 101.2, 102.2, 103.2, 104.2, 105.2, 106.2, 107.2, and 108.2, as well as other residue positions described herein, may be used as starting amino acid sequences for synthesizing other TdT polypeptides by subsequent rounds of evolution that incorporate new combinations of various amino acid differences from other polypeptides at residue positions that have remained unchanged throughout previous rounds of evolution. Further improvements may occur by including amino acid differences at residue positions that have remained unchanged throughout previous rounds of evolution.

[0130] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and the following: 14, 17, 19, 20, 21, 22, 24, 27, 28 / 304, 55, 66, 67, 68, 80, 99, 100, 110, 120, 130, 140, 150, 160, 170, 190, 200, 210, 220, 240, 270, 280, 304, 355, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 510, 520, 530, 540, 550, 560, 570, 580, 590, 610, 620, 630, 640, 650, 660, 670, 680, 700, 710, 720, 730, 740, 750, 760, 770, 780, 790, 800, 810, 820, 830, 840, 850, 860, 870, 880, 890, 9 and 373. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and has the following sequences compared to SEQ ID NO:4: 14G, 17R, 19S, 20F, 20G, 20S, 21G, 22D, 24E, 24Q, 24V, 27G, 27Y, 28S / 304L, 55I, 66A, 66E, 66L, 66R, 67K, 67R, 68R, 80S, 99S, 103R, 103S, 111R, 113Q, 115R, 140I ... 58R, 159L, 160S, 161S, 167E, 170Q, 180A, 180T, 192S, 197G, 200A, 200R, 201R, 219A, 219R, 219W, 233G, 233R, 235D, 235S, 235V, 238G, 246G, 249T, 256G, 258C, 267G, 268E, and amino acid sequences having one or more residue differences or sets of residue differences selected from 273R, 295D, 295S, 295V, 295W, 296V, 297A, 297L, 297V, 300S, 303G, 304E, 304W, 306L, 322V, 350V, 353N, 367C, and 373D.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4, and has at least one of the following sequences compared to SEQ ID NO: 4: P14G, T17R, E1 9S, L20F, L20G, L20S, E21G, V22D, S24E, S24Q, S24V, A27G, A27Y, R28S / S304L, M55I, I66A, I66E, I66L, I66R, F67K, F67R, C68R, Y80S, D99S, D103R, D103S, E111R, E113Q, S115R, V140I, K158R, A15 9L, D160S, K161S, K167E, K170Q, V180A, V180T, T192S, N197G, S200A, S200R, T201R, K219A, K219 R, K219W, P233G, P233R, E235D, E235S, E235V, E238G, L246G, K249T, C256G, I258C, Q267G, L268E, and an amino acid sequence having one or more residue differences or a set of residue differences selected from I273R, N295D, N295S, N295V, N295W, S296V, S297A, S297L, S297V, T300S, K303G, S304E, S304W, M306L, T322V, H350V, K353N, R367C, and G373D.

[0131] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and the following: 10, 17, 20, 27, 28 , 55, 62, 63, 66, 67, 68, 68 / 118, 87, 103, 106, 111, 131, 155, 157, 160, 160 / 296, 177, 181, 200, 219, 246, 256, 263, 292, 295, 296, 297, 315, and 373. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and has at least one of the following sequences compared to SEQ ID NO:4: 10Q, 17D, 20S, 27G, 28F, 28S, 55L, 62M, 63G, 66V, 67A, 67D, 68A, 68B, 68C, 68D, 68E, 68F, 68F, 68F, 68G, 68H, 68H, 68I, 68I, 68J, 68K ... and amino acid sequences having one or more residue differences or sets of residue differences selected from 67N, 68R, 68R / 118H, 87S, 103H, 106R, 111T, 131E, 155S, 157A, 157I, 160C, 160N / 296W, 160T, 177S, 181R, 200A, 219P, 246C, 256E, 263A, 292S, 295V, 296G, 297T, 315G, 315V, 373D, and 373R.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and has one of the following sequences compared to SEQ ID NO:4: S10Q, T17D, L20S, A27G, R28F, R28S, M55L, K62M, E63G, I66V, F67A, F67D, F67N, C68R, The amino acid sequence has one or more residue differences or a set of residue differences selected from C68R / K118H, R87S, D103H, E106R, E111T, Q131E, E155S, L157A, L157I, D160C, D160N / S296W, D160T, E177S, S181R, S200A, K219P, L246C, C256E, F263A, G292S, N295V, S296G, S297T, I315G, I315V, G373D, and G373R.

[0132] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and the following: 8, 10, 11, 15, 16, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and the following: 8, 10, 11, 15, 16, 20, 21, 22, 23, 24, 25, 26, 2, 24, 26, 28, 30, 34, 42, 44, 47, 48, 52, 54, 55, 62, 63, 66, 67, 68, 72, 77, 80, 84, 87, 89, 91, 92, 99, 101, 102, 103, 106, 109, 111, 111 / 346, 113, 115, 116, 118, 122, 123, 131, 140, 144, 147, 156, 157, 160, 160 / 296, 161, 162, 163, 164, 170, 173, 174, 175, 180, 181, 185, 189, 192, 193, 194, 197, 198, 198 / 289, 200, 201, 207, 219, 230, 234, 235, 237, 238, 241, 245, 246, 249, 256, 257, 258, 262, 263, 264, 267, 268, 272, and amino acid sequences having one or more residue differences or sets of residue differences selected from 273, 280, 284, 286, 289, 290, 291, 292, 295, 296, 297, 299, 300, 302, 304, 307, 308, 310, 311, 321, 322, 325, 350, 353, 355, 365, 366, 368, 371, 373, and 388. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4, and has one of the following sequences compared to SEQ ID NO: 4: 8P, 8R, 10D, 10G, 10H ... 0Q, 11R, 15F, 15G, 16V, 20G, 21G, 22G, 24N, 24T, 24Y, 26L, 28S, 30G, 30P, 34H, 42T, 44D, 47I, 48N, 52 L, 54L, 55L, 62L, 62M, 63G, 66A, 66E, 66R, 66V, 67A, 67D, 67G, 67K, 67N, 67R, 68R, 68S, 68T, 72D, 77V,80C, 80G, 80R, 80S, 80T, 84T, 87S, 89A, 91L, 92D, 92M, 92S, 99P, 101T, 102W, 103A, 103E, 103L, 103Q, 103S, 106R, 109C, 109L, 111A, 111I, 111M, 111P / 346H, 111R, 111S, 111T, 113C, 115E, 115R, 116I, 116P, 118M, 122S, 123G, 131L, 140I, 144R, 147L, 147S, 147Y, 156 F,157A,157I,157V,160C,160N / 296W,160T,161L,161R,162R,163R,164D,164R,170D,173L,174R,175D,180A,180R,180T,181R,1 85R, 189A, 192Y, 193A, 193G, 193W, 194K, 197S, 198G, 198G / 289H, 200C, 200N, 201L, 201N, 207G, 219G, 219P, 230C, 234L, 235D, 235Q , 235S, 237R, 238L, 238S, 238Y, 241D, 245E, 246C, 246V, 249Q, 256E, 256G, 256P, 257F, 258S, 262G, 262R, 263A, 263K, 264T, 264Y, 2 67D, 267S, 267V, 267W, 268V, 272G, 273P, 273Q, 280C, 284F, 286A, 289G, 289R, 290K, 290R, 291K, 291Q, 292S, 292V, 295A, 296R, 296W , 296Y, 297P, 297T, 299V, 300L, 300R, 300S, 302A, 304A, 304K, 307I, 308M, 310G, 311V, 321E, 321G, 322R, 322V, 325L, 350E, 350V, 350Y, 353A, 353F, 353N, 355E, 355F, 365D, 365R, 366P, 368L, 371I, 373N, and 388Q. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4,and the following relative to SEQ ID NO: 4: G8P, G8R, S10D, S10G, S10Q, G11R, S15F, S15G, N16V, L20G, E21G, V22G, S24N, S24T, S24Y, I26L, R28S, V30G, V30P, S34H, N42T, Y44D, K47I, F48N, F52L, I54L, M55L, K62L, K62M, E63G, I66A, I66E, I66R, I66V, F67A, F67D, F67G, F67K, F67N, F67R, C68R, C68S, C68T, L72D, L77V, Y80C, Y80G, Y 80R, Y80S, Y80T, P84T, R87S, K89A, I91L, Q92D, Q92M, Q92S, D99P, V101T, R1 02W, D103A, D103E, D103L, D103Q, D103S, E106R, I109C, I109L, E111A, E111I , E111M, E111P / R346H, E111R, E111S, E111T, E113C, S115E, S115R, R116I, R 116P, K118M, N122S, D123G, Q131L, V140I, E144R, F147L, F147S, F147Y, E156 F, L157A, L157I, L157V, D160C, D160N / S296W, D160T, K161L, K161R, S162R, L163R, K164D, K164R, K170D, F173L, L174R, Y175D, V180A, V180R, V180T, S18 1R, K185R, D189A, T192Y, L193A, L193G, L193W, I194K, N197S, T198G, T198G / Y289H, S200C, S200N, T201L, T201N, L207G, K219G, K219P, N230C, K234L, E2 35D, E235Q, E235S, D237R, E238L, E238S, E238Y, H241D, D245E, L246C, L246 V, K249Q, C256E, C256G, C256P, D257F, I258S, T262G, T262R, F263A, F263K, V 264T, V264Y, Q267D, Q267S, Q267V, Q267W, L268V, K272G, I273P, I273Q, Q28 0C, A284F, L286A, Y289G, Y289R, Q290K, Q290R, P291K, P291Q, G292S, G292V,The amino acid sequence may have one or more residue differences or a set of residue differences selected from N295A, S296R, S296W, S296Y, S297P, S297T, N299V, T300L, T300R, T300S, K302A, S304A, S304K, A307I, E308M, K310G, D311V, I321E, I321G, T322R, T322V, E325L, H350E, H350V, H350Y, K353A, K353F, K353N, I355E, I355F, K365D, K365R, K366P, I368L, K371I, G373N, and E388Q.

[0133] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and one or more residue differences or sets of residue differences selected from the following compared to SEQ ID NO:4: 60, 60 / 259, 60 / 278, and 65 / 259. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and has one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO:4: 60K, 60K / 259K, 60K / 278H, and 65K / 259Q. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and has one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO:4: E60K, E60K / I259K, E60K / N278H, and E65K / I259Q.

[0134] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and the following sequences compared to SEQ ID NO:4: 20, 20 / 21 / 68 / 103 / 200 / 238 / 297, 20 / 21 / 68 / 111 / 235, 20 / 21 / 68 / 160, 20 / 21 / 68 / 160 / 180 / 200 / 246, 20 / 21 / 68 / 160 / 246, 20 / 21 / 68 / 180, 20 / 21 / 68 / 180 / 235, 20 / 21 / 68 / 200 / 235 / 297, 20 / 21 / 68 / 233 / 246 / 297, 20 / 21 / 103 / 233, 20 / 21 / 111, 20 / 21 / 111 / 160 / 200, 20 / 21 / 111 / 200 / 238, 20 / 21 / 180 / 297, 20 / 21 / 233, 20 / 68 / 103 / 160 / 200, 20 / 68 / 103 / 180 / 200, 20 / 68 / 160, 20 / 68 / 160 / 180, 20 / 68 / 160 / 180 / 23 3 / 235 / 246, 20 / 68 / 160 / 235 / 297, 20 / 68 / 160 / 246, 20 / 68 / 200 / 238 / 246, 20 / 68 / 235 / 297, 20 / 103 / 160 / 180 / 200 / 235, 20 / 103 / 233, 20 / 111, 20 / 111 / 180 / 235 / 246 / 297, 21, 21 / 68, 21 / 68 / 103 / 111, 21 / 68 / 111 / 200, 21 / 68 / 160 / 180 / 200 / 205 / 297, 21 / 68 / 160 / 180 / 200 / 297, 21 / 68 / 160 / 238, 21 / 68 / 160 / 238 / 246, 21 / 68 / 180, 21 / 68 / 180 / 235, 21 / 68 / 180 / 246, 21 / 68 / 200, 21 / 68 / 235, 21 / 103 / 233, 21 / 233 / 297, 68, 68 / 103 / 160 / 235, 68 / 103 / 200 / 235 / 246 / 297, 68 / 111 / 200 / 238, 68 / 111 / 233 / 236 / 297, 68 / 160 / 233 / 246, 68 / 200 / 235 / 297, 103, 103 / 160 / 180, 103 / 160 / 297, 103 / 233, 111,and 111 / 160 / 233 / 235 / 297. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and the following compared to SEQ ID NO:4: 20G / 21G / 68R / 103S / 200R / 238G / 297A, 20G / 21G / 68R / 160S / 180A / 200R / 246G, 20G / 21G / 68R / 180A, 20G / 21G / 68R / 180A / 235S, 20G / 21G / 111R / 160S / 200A, 20G / 21G / 180A / 297A , 20G / 21G / 233G, 20G / 68R / 103S / 160S / 200R, 20G / 68R / 160S, 20S, 20S / 21G / 68R / 111R / 235S, 20S / 21G / 68R / 160S, 20S / 21G / 68R / 160S / 246G, 2 0S / 21G / 68R / 200R / 235S / 297A, 20S / 21G / 68R / 233G / 246G / 297A, 20S / 21 G / 103S / 233G, 20S / 21G / 111R, 20S / 21G / 111R / 200R / 238G, 20S / 68R / 103S / 180A / 200A, 20S / 68R / 160S / 180A, 20S / 68R / 160S / 180A / 233G / 235S / 246G, 20S / 68R / 160S / 235S / 297A, 20S / 68R / 160S / 246G, 20S / 68R / 200 A / 238G / 246G, 20S / 68R / 235S / 297A, 20S / 103S / 160S / 180A / 200R / 235S, 20S / 103S / 233G, 20S / 111R, 20S / 111R / 180A / 235S / 246G / 297A, 21G, 21 G / 68R, 21G / 68R / 103S / 111R, 21G / 68R / 111R / 200R, 21G / 68R / 160S / 180 A / 200R / 205A / 297A, 21G / 68R / 160S / 180A / 200R / 297A, 21G / 68R / 160S / 238G, 21G / 68R / 160S / 238G / 246G, 21G / 68R / 180A, 21G / 68R / 180A / 235S,21G / 68R / 180A / 246G, 21G / 68R / 200R, 21G / 68R / 235S, 21G / 103S / 233G, 21G / 233G / 297A, 68R, 68R / 1 03S / 160S / 235S, 68R / 103S / 200R / 235S / 246G / 297A, 68R / 111R / 200A / 238G, 68R / 111R / 233G / 236N / and amino acid sequences having one or more residue differences or sets of residue differences selected from 297A, 68R / 160S / 233G / 246G, 68R / 200A / 235S / 297A, 103S, 103S / 160S / 180A, 103S / 160S / 297A, 103S / 233G, 111R, and 111R / 160S / 233G / 235S / 297A. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and the following sequence identity compared to SEQ ID NO:4: L20G / E21G / C68R / D103S / S200R / E238G / S297A, L20G / E21G / C68R / D160S / V180A / S200R / L246G, L20G / E21G / C68R / V 180A, L20G / E21G / C68R / V180A / E235S, L20G / E21G / E111R / D160S / S200A, L20G / E21G / V180A / S297A , L20G / E21G / P233G, L20G / C68R / D103S / D160S / S200R, L20G / C68R / D160S, L20S, L20S / E21G / C68R / E111R / E235S, L20S / E21G / C68R / D160S, L20S / E21G / C68R / D160S / L246G, L20S / E21G / C68R / S200R / E235S / S297A, L20S / E21G / C68R / P233G / L246G / S297A, L20S / E21G / D103S / P233G, L20S / E21G / E11 1R, L20S / E21G / E111R / S200R / E238G, L20S / C68R / D103S / V180A / S200A, L20S / C68R / D160S / V180A,L20S / C68R / D160S / V180A / P233G / E235S / L246G, L20S / C68R / D160S / E235S / S297A, L20S / C68R / D160S / L246G, L20S / C68R / S200A / E238G / L246G, L20S / C68R / E235S / S297A, L20S / D103S / D160S / V180A / S200R / E235S, L20S / D103S / P233G, L20S / E111R, L20S / E1 11R / V180A / E235S / L246G / S297A, E21G, E21G / C68R, E21G / C68R / D103S / E111R, E21G / C68R / E111R / S200R, E21G / C68R / D160S / V1 80A / S200R / D205A / S297A, E21G / C68R / D160S / V180A / S200R / S297A, E21G / C68R / D160S / E238G, E21G / C68R / D160S / E238G / L246G, E21G / C68R / V180A, E21G / C68R / V180A / E235S, E21G / C68R / V180A / L246G, E21G / C68R / S200R, E21G / C68R / E235S, E21G / D103S / P2 33G, E21G / P233G / S297A, C68R, C68R / D103S / D160S / E235S, C68R / D103S / S200R / E235S / L246G / S297A, C68R / E111R / S200A / E238G , C68R / E111R / P233G / D236N / S297A, C68R / D160S / P233G / L246G, C68R / S200A / E235S / S297A, D103S, D103S / D160S / V180A, D103S / D160S / S297A, D103S / P233G, E111R, and E111R / D160S / P233G / E235S / S297A.

[0135] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 580, and the following sequences compared to SEQ ID NO: 580: 55, 55 / 67, 55 / 67 / 87, 55 / 67 / 87 / 106 / 111 / 315, 55 / 67 / 87 / 157 / 315, 55 / 67 / 106 / 111 / 157, 55 / 67 / 106 / 315, 55 / 67 / 111, 55 / 67 / 111 / 315, 55 / 67 / 315, 55 / 87, 55 / 87 / 106, 55 / 87 / 106 / 111 / 315, 55 / 87 / 106 / 315, 55 / 87 / 157, 55 / 87 / 157 / 207, 55 / 106 / 111, 55 / 106 / 157, 55 / 111, 55 / 111 / 315, 55 / 157, 55 / 315, 67, 67 / 87, 67 / 87 / 106 / 157, 67 / 87 / 111 / 157 / 315, 67 / 87 / 157, 67 / 87 / 315, 67 / 106, 67 / 106 / 111, 67 / 106 / 111 / 315, 67 / 106 / 157, 67 / 111, 67 / 111 / 315, 67 / 157, 67 / 157 / 315, 87, 87 / 106, 87 / 106 / 111 / 315, 87 / 111 / 157 / 315, 87 / 157, 87 / 157 / 315, 87 / 315, 106, 106 / 111 / 157, 106 / 315, 157, 157 / 315, and 315. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:580, and The following: 55L, 55L / 67A / 106R / 111T / 157I, 55L / 67A / 111T, 55L / 67A / 111T / 315V, 55L / 67D, 55L / 67D / 87S / 157I / 315V, 55L / 67D / 315V, 55L / 67M / 87S, 55L / 67M / 106R / 315V, 55L / 67N / 87S,55L / 67N / 87S / 106R / 111T / 315V, 55L / 87S, 55L / 87S / 106R, 55L / 87S / 106R / 111T / 315V, 55L / 87S / 1 06R / 315V, 55L / 87S / 157A, 55L / 87S / 157I, 55L / 87S / 157I / 207Q, 55L / 106R / 111T, 55L / 106R / 157I, 55L / 111T, 55L / 111T / 315V, 55L / 157I, 55L / 315V, 67A, 67A / 87S / 106R / 157A, 67A / 87S / 111T / 157A / 315V, 67A / 106R / 111T / 315V, 67A / 111T, 67A / 157I, 67D, 67D / 111T, 67M, 67M / 87S, 67M / 87S / 157A, 67M / 87S / 157I, 67M / 106R / 157A, 67M / 111T / 315V, 67M / 157A, 67M / 157A / 315V, 67N, 67N / 87S, 67N / 87S / 315V, 67N / 106R, 67N / 106R / 111T, 87S, 87S / 106R, 87S / 106R / 111T / 315V, 87S / 111T / 157A / 315 and 315V, 87S / 111T / 157I / 315V, 87S / 157A, 87S / 157I, 87S / 157I / 315V, 87S / 315V, 106R, 106R / 111T / 157I, 106R / 315V, 157A, 157I, 157I / 315V, and 315V. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 580, and has at least one of the following sequences compared to SEQ ID NO: 580: M55L, M55L / F6 7A / E106R / E111T / L157I, M55L / F67A / E111T, M55L / F67A / E111T / I315V, M55L / F67D, M55L / F67D / R87 S / L157I / I315V, M55L / F67D / I315V, M55L / F67M / R87S, M55L / F67M / E106R / I315V, M55L / F67N / R87S,M55L / F67N / R87S / E106R / E111T / I315V, M55L / R87S, M55L / R87S / E106R, M55L / R87S / E106R / E111T / I315V, M55L / R87S / E106R / I315V, M55L / R87S / L157A, M55L / R87S / L157I, M55L / R87S / L157I / L207Q, M55L / E106R / E111T, M55L / E106R / L15 7I, M55L / E111T, M55L / E111T / I315V, M55L / L157I, M55L / I315V, F67A, F67A / R87S / E106R / L157A, F67A / R87S / E111T / L 157A / I315V, F67A / E106R / E111T / I315V, F67A / E111T, F67A / L157I, F67D, F67D / E111T, F67M, F67M / R87S, F67M / R87S / L157A, F67M / R87S / L157I, F67M / E106R / L157A, F67M / E111T / I315V, F67M / L157A, F67M / L157A / I315V, F67N, F67N / R87 S, F67N / R87S / I315V, F67N / E106R, F67N / E106R / E111T, R87S, R87S / E106R, R87S / E106R / E111T / I315V, R87S / E111T / L and I315V, R87S / L157A / I315V, R87S / E111T / L157I / I315V, R87S / L157A, R87S / L157I, R87S / L157I / I315V, R87S / I315V, E106R, E106R / E111T / L157I, E106R / I315V, L157A, L157I, L157I / I315V, and I315V.

[0136] In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 580, and the following sequences compared to SEQ ID NO: 580: 59 / 62 / 63 / 68, 59 / 62 / 63 / 68 / 103 / 234, 59 / 62 / 63 / 68 / 147, 60 / 62 / 68 / 91 / 111 / 234 / and 147 / 210 / 234. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 580, and has at least one of the following sequences compared to SEQ ID NO: 580: 59A / 62N / 63G / 68C, 59A / 62N / 63G / 68C / 103R / 234G, 59A / 62N / 63G / 68C / 147Y, 60V / 62N / 68C / 91L / 111Y / 234G / 289P, 62N / 63G / 68C, 60V / ... and 147Y / 210L / 234G.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 580, and has at least one of the following sequences compared to SEQ ID NO: 580: Y59A / K62N / E63G / R68C, Y59A / K62N / E63G / R68C / D103R / K234G, Y59A / K62N / E63G / R68C / F147Y, E60V / K62N / R68C / I91L / E111Y / K234G / Y289P, K62N / E63G / R68C ... 3G / R68C / I91L / I109L / I210L, K62N / E63G / R68C / I91L / F147Y / D205G / I210L / K234G, K62 N / R68C / I91L / E111Y / Y289P, K62N / R68C / D103R, K62N / R68C / E111Y, I91L, I91L / I109L / E and an amino acid sequence having one or more residue differences or a set of residue differences selected from: 111Y, I91L / I109L / F147Y / D205G / I210L / K234G, I109L / E111Y / D205G / I210L / K234G / Y289P, E111Y, and F147Y / I210L / K234G.

[0137] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 692, and the following sequences compared to SEQ ID NO: 692: 52 / 55 / 106 / 256, 52 / 55 / 181 / 235 / 256, 52 / 55 / 181 / 256, 52 / 9 and amino acid sequences having one or more residue differences or sets of residue differences selected from: 9 / 181 / 235, 52 / 106 / 181 / 235 / 256, 52 / 106 / 235 / 256, 52 / 106 / 256, 52 / 173 / 235, 52 / 235, 52 / 235 / 256, 55 / 99 / 181 / 256, 55 / 99 / 256 / 350, 55 / 181 / 235 / 256 / 350, 55 / 256, 99 / 235, 106, 173, and 235. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 692, and has at least one of the following sequences compared to SEQ ID NO: 692: 52L / 55L / 106R / 256G, 52L / 55L / 181R / 235Q / 256E, 52L / 55L / 181R / 256G, 52L / 99P / 181R / 235Q ... and amino acid sequences having one or more residue differences or sets of residue differences selected from L / 106R / 181R / 235Q / 256G, 52L / 106R / 235Q / 256G, 52L / 106R / 256E, 52L / 173L / 235Q, 52L / 235Q, 52L / 235Q / 256E, 55L / 99P / 181R / 256G, 55L / 99P / 256G / 350V, 55L / 181R / 235Q / 256E / 350V, 55L / 256G, 99P / 235Q, 106R, 173L, and 235Q.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 692, and has at least one of the following sequences compared to SEQ ID NO: 692: F52L / M55L / E106R / C256G, F52L / M55L / S181R / E235Q / C256E, F52L / M55L / S181R / C256G, F52L / D99P / S181R / E235Q, F52L / E106R / S1 and E235Q.

[0138] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and the following sequences compared to SEQ ID NO:4: 20 / 21 / 52 / 67 / 68 / 87 / 111 / 157 / 160 / 173 / 180 / 200 / 235 / 246 / 315, 20 / 21 / 55 / 67 / 68 / 87 / 111 / 157 / 160 / 180 / 181 ... and 20 / 21 / 68 / 160 / 180 / 200 / 246 / 256 / 315, 20 / 21 / 60 / 72 / 160 / 180 / 200 / 246 / 259 / 338 / 358, 20 / 21 / 67 / 68 / 87 / 111 / 157 / 160 / 180 / 200 / 235 / 246 / 315, 20 / 21 / 67 / 68 / 87 / 111 / 157 / 160 / 180 / 200 / 246 / 315, and 20 / 21 / 68 / 160 / 180 / 200 / 246.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and has at least 20G / 21G / 52L / 67A / 68R / 87S / 111T / 157A / 160S / 173L / 180A / 200R / 235Q / 246G / 315V, 20G / 21G / 5 5L / 67A / 68R / 87S / 111T / 157A / 160S / 180A / 181R / 200R / 235Q / 246G / 256E / 315V / 350V, 20G / 21 G / 55L / 67A / 68R / 87S / 111T / 157A / 160S / 180A / 200R / 246G / 256G / 315V, 20G / 21G / 60K / 72K / 1 60S / 180A / 200R / 246G / 259K / 338K / 358R, 20G / 21G / 67A / 68R / 87S / 111T / 157A / 160S / 180A / 2 and 20G / 21G / 68R / 160S / 180A / 200R / 246G / 315V.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4, and has one of the following sequences compared to SEQ ID NO:4: L20G / E21G / F52L / F67A / C68R / R87S / E111T / L157A / D160S / F173L / V180A / S200R / E235Q / L246G / I315, L20G / E21G / M55L / F67A / C68R / R87 S / E111T / L157A / D160S / V180A / S181R / S200R / E235Q / L246G / C256E / I315V / H350V, L20G / E21G / M55L / F67A / C68R / R87S / E111T / L157A / D160S / V180A / S200R / L246G / C256G / I315V, L20G / E21G / E60K / L72K / D160S / V 180A / S200R / L246G / I259K / R338K / N358R, L20G / E21G / F67A / C68R / R87S / E111T / L157A / D160S / V180A / S2 and L20G / E21G / C68R / D160S / V180A / S200R / L246G / I315V, L20G / E21G / F67A / C68R / R87S / E111T / L157A / D160S / V180A / S200R / L246G / I315V, and L20G / E21G / C68R / D160S / V180A / S200R / L246G.

[0139] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 882, and one or more residue differences or sets of residue differences selected from the following compared to SEQ ID NO: 882: 63, 68, 72, 211, 220, 275, 349, and 350. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 882 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 882: 63G, 68M, 72R, 211C, 220R, 275R, 349E, 349M, 349R, 350E, and 350Y. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 882 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 882: E63G, C68M, K72R, T211C, M220R, A275R, S349E, S349M, S349R, H350E, and H350Y.

[0140] In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 882, and the following compared to SEQ ID NO: 882: 20 / 21, 20 / 21 / 67, 20 / 21 / 67 / 87, 20 / 21 / 67 / 87 / 111, 20 / 21 / 67 / 87 / 111 / 200, 20 / 21 / 67 / 87 / 111 / 315, 20 / 21 / 67 / 8 7 / 157 / 180, 20 / 21 / 67 / 87 / 246 / 315, 20 / 21 / 67 / 160, 20 / 21 / 87 / 111 / 246 / 315, 20 / 21 / 87 / 157 / 200, 20 / 21 / 87 / 180 / 246 / 315, 20 / 21 / 87 / 315, 20 / 21 / 1 11 / 157 / 315, 20 / 21 / 111 / 246, 20 / 21 / 315, 20 / 67 / 87 / 111 / 180, 20 / 67 / 87 / 157 / 160 / 180 / 315, 20 / 68 / 111 / 157 / 160, 20 / 68 / 160 / 200 / 246, 20 / 87 / 111 / 180, 20 / 87 / 157 / 160, 20 / 160 / 315, 20 / 200, 21, 21 / 67 / 87 / 111 / 200, 21 / 67 / 87 / 157 / 246, 21 / 67 / 180 / 315, 21 / 67 / 200, 21 / 87 / 157 / 160, 21 / 87 / 160 / 2 00 / 315, 21 / 87 / 160 / 315, 21 / 87 / 200, 21 / 87 / 246 / 315, 21 / 111, 21 / 157 / 160, 21 / 160 / 315, 21 / 200 / 315, 21 / 246, 67, 67 / 87, 67 / 87 / 111 / 157 / 160 / 315, In some embodiments, the amino acid sequence has one or more residue differences or a set of residue differences selected from: 67 / 111, 67 / 157 / 160 / 180, 67 / 157 / 160 / 315, 67 / 160, 67 / 180 / 200, 67 / 180 / 200 / 315, 67 / 315, 68 / 87, 68 / 157 / 160 / 200 / 315, 87, 87 / 111, 87 / 111 / 200, 87 / 111 / 200 / 246, 87 / 157, 87 / 157 / 180 / 200, 111 / 157 / 180 / 200 / 315, 157, 157 / 160, 246, and 315.Engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher sequence identity to the reference sequence SEQ ID NO: 882, and include the following sequences compared to SEQ ID NO: 882: 20L / 21E, 20L / 21E / 67A, 20L / 21E / 67A / 87S, 20L / 21E / 67A / 87S / 111T, 20L / 21E / 67A / 87S / 111T / 200S, 20L / 21E / 67A / 87S / 111T / 315V, 20L / 21E / 67A / 87S / 157A / 180V, 20L / 21E / 67A / 87S / 246L / 315V, 20L / 21E / 67 A / 160D, 20L / 21E / 87S / 111T / 246L / 315V, 20L / 21E / 87S / 157A / 200S, 20L / 21E / 87S / 180V / 246L / 315V, 20L / 21E / 87S / 315V, 20L / 21E / 111T / 157A / 315V, 20 L / 21E / 111T / 246L, 20L / 21E / 315V, 20L / 67A / 87S / 111T / 180V, 20L / 67A / 87S / 157A / 160D / 180V / 315V, 20L / 68R / 111T / 157A / 160D, 20L / 68R / 160D / 200S / 2 46L, 20L / 87S / 111T / 180V, 20L / 87S / 157A / 160D, 20L / 160D / 315V, 20L / 200S, 21E, 21E / 67A / 87S / 111T / 200S, 21E / 67A / 87S / 157A / 246L, 21E / 67A / 180V / 3 15V, 21E / 67A / 200S, 21E / 87S / 157A / 160D, 21E / 87S / 160D / 200S / 315V, 21E / 8 7S / 160D / 315V, 21E / 87S / 200S, 21E / 87S / 246L / 315V, 21E / 111T, 21E / 157A / 160D, 21E / 160D / 315V, 21E / 200S / 315V, 21E / 246L, 67A, 67A / 87S, 67A / 87S / 1 11T / 157A / 160D / 315V, 67A / 111T, 67A / 157A / 160D / 180V, 67A / 157A / 160D / 3 15V, 67A / 160D, 67A / 180V / 200S, 67A / 180V / 200S / 315V, 67A / 315V, 68R / 87S,and amino acid sequences having one or more residue differences or sets of residue differences selected from 68R / 157A / 160D / 200S / 315V, 87S, 87S / 111T, 87S / 111T / 200S, 87S / 111T / 200S / 246L, 87S / 157A, 87S / 157A / 180V / 200S, 111T / 157A / 180V / 200S / 315V, 157A, 157A / 160D, 246L, and 315V. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 882, and has at least one of the following sequence identity compared to SEQ ID NO: 882: G20L / G21E, G20L / G21E / F67A, G20L / G21E / F67A / R87S ... G21E / F67A / R87S / E111T, G20L / G21E / F67A / R87S / E111T / R200S, G20L / G21E / F67A / R87S / E111T / I315V, G20L / G21E / F67A / R87S / L157A / A180V, G20L / G21E / F67A / R87S / G246L / I315V, G20L / G21E / F67A / S160D, G20L / G21E / R87S / E111T / G246L / I315 V, G20L / G21E / R87S / L157A / R200S, G20L / G21E / R87S / A180V / G246L / I315V, G20L / G21E / R87S / I315V, G20L / G21E / E111T / L 157A / I315V, G20L / G21E / E111T / G246L, G20L / G21E / I315V, G20L / F67A / R87S / E111T / A180V, G20L / F67A / R87S / L157A / S160 D / A180V / I315V, G20L / C68R / E111T / L157A / S160D, G20L / C68R / S160D / R200S / G246L, G20L / R87S / E111T / A180V, G20L / R87 S / L157A / S160D, G20L / S160D / I315V, G20L / R200S, G21E, G21E / F67A / R87S / E111T / R200S, G21E / F67A / R87S / L157A / G246L,G21E / F67A / A180V / I315V, G21E / F67A / R200S, G21E / R87S / L157A / S160D, G21E / R87S / S160D / R 200S / I315V, G21E / R87S / S160D / I315V, G21E / R87S / R200S, G21E / R87S / G246L / I315V, G21E / E1 11T, G21E / L157A / S160D, G21E / S160D / I315V, G21E / R200S / I315V, G21E / G246L, F67A, F67A / R8 7S, F67A / R87S / E111T / L157A / S160D / I315V, F67A / E111T, F67A / L157A / S160D / A180V, F67A / L1 57A / S160D / I315V, F67A / S160D, F67A / A180V / R200S, F67A / A180V / R200S / I315V, F67A / I315V, C68R / R87S, C68R / L157A / S160D / R200S / I315V, R87S, R87S / E111T, R87S / E111T / R200S, R87S / E and amino acid sequences having one or more residue differences or sets of residue differences selected from: 111T / R200S / G246L, R87S / L157A, R87S / L157A / A180V / R200S, E111T / L157A / A180V / R200S / I315V, L157A, L157A / S160D, G246L, and I315V.

[0141] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 914, and the following: 52, 52 / 180, 52 / 180 / 200 compared to SEQ ID NO: 914. 52 / 200 / 349, 52 / 200 / 315, 52 / 200 / 349, 52 / 200 / 349 / 350, 52 / 315, 52 / 315 / 349, 52 / 349, 52 / 349 / 350, 180, and 349. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 914, and has at least one of the following sequences compared to SEQ ID NO: 914: 52L, 52L / 180V, 52L / 180V / 200S / 235Q / 315V / 349 and 52L / 180V / 200S / 349E, 52L / 180V / 349E, 52L / 200S, 52L / 200S / 315V, 52L / 200S / 349E, 52L / 200S / 349E / 350E, 52L / 315V, 52L / 315V / 349E, 52L / 349E, 52L / 349E / 350E, 180V, and 349E.In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 914, and at least one of the following sequences compared to SEQ ID NO: 914: F52L, F52L / A180V, F52L / A180V / R200S / E235Q / I315V / S349E ... and amino acid sequences having one or more residue differences or sets of residue differences selected from V / R200S / S349E, F52L / A180V / S349E, F52L / R200S, F52L / R200S / I315V, F52L / R200S / S349E, F52L / R200S / S349E / H350E, F52L / I315V, F52L / I315V / S349E, F52L / S349E, F52L / S349E / H350E, A180V, and S349E.

[0142] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1034, and the following sequences compared to SEQ ID NO: 1034: 53 / 219 / 358, 55, 55 / 99 / 103, 55 / 99 / 219 / 358 / 373, 55 / 103 / 181, 55 / 103 / 219, 55 / 103 / 338, 55 / 181 / 219, 55 / 181 / 246 ... and 358.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:1034, and Compare to: 53Q / 219P / 358N, 55L, 55L / 99P / 103A, 55L / 99P / 219P / 358N / 373D, 55L / 103A / 181R, 55L / 103A / 219P, 55L / 103A / 338R, 55L / 181R / 219P, 55L / 181R / 246G, 55L / 181R / 358N, 55 L / 219P / 246G / 358N, 55L / 219P / 256G / 338R, 55L / 256G / 259I, 99P / 103A / 219P, 103A / 219P / 256E, 103A / 219P / 338R / 358N, 103A / 256G, 103A / 259I, 106R / 173L / 200S / 235Q / 315V, 106 and amino acid sequences having one or more residue differences or sets of residue differences selected from R / 173L / 388Q, 111T / 173L, 111T / 173L / 235Q / 315V, 173L, 173L / 297A, 181R, 219P, 219P / 358N, 256E, 256G, 256G / 259I, and 358N.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1034, and the following: E53Q / K219P / R35 compared to SEQ ID NO: 1034: 8N, M55L, M55L / D99P / D103A, M55L / D99P / K219P / R358N / G373D, M55L / D103A / S181R, M55L / D103A / K21 9P, M55L / D103A / K338R, M55L / S181R / K219P, M55L / S181R / L246G, M55L / S181R / R358N, M55L / K219P / L2 46G / R358N, M55L / K219P / C256G / K338R, M55L / C256G / K259I, D99P / D103A / K219P, D103A / K219P / C256 E, D103A / K219P / K338R / R358N, D103A / C256G, D103A / K259I, E106R / F173L / R200S / E235Q / I315V, E106 and amino acid sequences having one or more residue differences or sets of residue differences selected from R / F173L / E388Q, E111T / F173L, E111T / F173L / E235Q / I315V, F173L, F173L / S297A, S181R, K219P, K219P / R358N, C256E, C256G, C256G / K259I, and R358N.

[0143] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1034, and the following: 48, 52, 53, 219, 358, 55, 55 / 9 and amino acid sequences having one or more residue differences or sets of residue differences selected from: 9 / 219 / 358 / 373, 55 / 181 / 358, 55 / 219 / 246 / 358, 55 / 256, 56, 58, 62, 65, 66, 70, 71, 74, 77, 78, 79, 103 / 219 / 338 / 358, 219, 219 / 358, 328, 345, 350, 353, 355, and 358. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1034, and has at least one of the following sequences compared to SEQ ID NO: 1034: 48T, 52A, 53Q / 219P / 358N, 55L, 55L / 99P / 219P / 358N / 373D, 55L / 181R / 358N, 55L / 219P / 246G / 358N, 55L / 256E, 56V, 58D, 62G, 65L, 66P, 66S, 70V, 71A, 74G, 77I, 77V, 78F, 79R, 103A / 219P / 338R / 358N, 219L, 219P / 358N, 219V, 328F, 345V, 350T, 353S, 355V, and 358N.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1034, and at least one of the following sequences compared to SEQ ID NO: 1034: F48T, L52A, E53Q / K219P / R358N, M55L, M55L / D99P / K219P / R358N / G373D, M55L / S181R / R358N , M55L / K219P / L246G / R358N, M55L / C256E, A56V, N58D, K62G, E65L, I66P, I66S, E70V, F71A, A74G, L77I, L77V, L78F, K79R, D103A / K219P / K338R / R358N, K219L, K219P / R358N, K219V, A328F, R345V, E350T, K353S, I355V, and R358N.

[0144] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1034, and the following: 47, 48, 51, 52 , 54, 55, 57, 58, 59, 61, 62, 63, 64, 65, 66, 69, 70, 71, 73, 74, 77, 79, 131, 133, 134, 219, 319, 321, 322, 325, 328, 332, 345, 350, 351, 352, 353, and 355. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1034, and has at least one of the following sequences compared to SEQ ID NO: 1034: 47Q, 48I, 48Y, 51L, 51S, 52A, 54N, 54V, 55R, 57C, 58C, 58D, 58M, 58S, 59W, 61T, 62G, 62R, 62W, 63D, 64Q, 64R, 65L, 66Q, 66R, 67Q, 67R, 68Q, 68R, 69Q, 70Q, 70Q, 71Q, 71Q, 72Q, 72Q, 73Q, 74Q, 75Q, 75Q, 76Q, 77Q, 78Q, 79Q, 79Q, 80Q, 81Q, 82Q, 83Q, 84Q, 85Q, 86Q, 87Q, 88Q, 89Q, 90Q, 91Q, 92Q, 93Q, 94Q, 95Q, 96Q, 97Q, 98Q, 9 ... and 355V.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1034, and any of the following sequences compared to SEQ ID NO: 1034: K47Q, F48I, F48Y, A51L, A51S, L52A, I54N, I54V, M55R, E57C, N58C, N58D, N58M, N58S, Y59W, F61T, K62G, K62R, K62W, E63D, N64Q, N64R, E65L, I66P, I66R, I66S, L69I, E70A, E70H, E70I, E70V, F71A, F71S, R73G, A74G, L77I, L77V, K79R, Q131 V, T133Y, S134M, S134V, K219L, K219V, L319F, I321V, T322S, E325S, A328F, A328P, A3 and an amino acid sequence having one or more residue differences or a set of residue differences selected from 28S, A328W, L332C, R345A, R345Q, R345V, E350S, E351Q, R352A, R352S, K353G, K353M, K353Q, I355A, I355L, and I355V.

[0145] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1270, and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 1270: 23, 32, 36, 40, 49, 93, 98, 107, 117, 124, 165, 182, 186, 269, 288, 309, 374, 377, and 386. In some embodiments, an engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1270, and has one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1270: 23L, 32H, 36H, 40S, 49S, 93Q, 93W, 98P, 107G, 107S, 117I, 124P, 165M, 182G, 186D, 186R, 269R, 288H, 309K, 374E, 374G, 377N, and 386V. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1270, and has at least one of the following sequences compared to SEQ ID NO: 1270: P23L, Q32H, Q and Y386V, 36H, T40S, T49S, G93Q, G93W, G98P, E107G, E107S, V117I, E124P, L165M, C182G, A186D, A186R, P269R, L288H, V309K, S374E, S374G, E377N, and Y386V.

[0146] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1270, and one or more residue differences or sets of residue differences selected from the following: 23, 40, 49, 107, 112, 120, 186, 271, 309, 374, and 377 compared to SEQ ID NO: 1270. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1270, and has one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1270: 23L, 40S, 49S, 107G, 107S, 112T, 120T, 186D, 271Q, 309K, 374E, and 377N. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1270 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1270: P23L, T40S, T49S, E107G, E107S, G112T, V120T, A186D, R271Q, V309K, S374E, and E377N.

[0147] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1270, and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 1270: 23, 182, 186, 269, 270, 293, 360, 372, and 378. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1270 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1270: 23V, 182A, 182R, 186E, 269A, 270T, 293G, 360A, 372G, and 378C. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1270, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1270: P23V, C182A, C182R, A186E, P269A, S270T, V293G, G360A, A372G, and I378C.

[0148] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and the following sequences compared to SEQ ID NO: 1344: 55 / 58 / 256 / 350 / 373 / 58 / 69 / and amino acid sequences having one or more residue differences or sets of residue differences selected from: 256 / 373, 58 / 99 / 351 / 355, 58 / 256, 70 / 71, 70 / 71 / 353, 70 / 134 / 353, 70 / 353, 71 / 77 / 353, 71 / 353, 77, 77 / 134 / 353, 77 / 353, 99 / 256 / 351 / 355, 134 / 353, 256, 350, and 353. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and has at least one of the following sequences compared to SEQ ID NO: 1344: 55R / 58D / 256E / 350S / 373D, 58D, 58S / 69I / 256E / 373D, 58S / 99V / and amino acid sequences having one or more residue differences or sets of residue differences selected from 351Q / 355A, 58S / 256E, 70V / 71A, 70V / 71A / 353G, 70V / 134M / 353G, 70V / 353M, 71A / 77I / 353G, 71A / 353G, 77I, 77I / 134V / 353G, 77I / 353M, 99V / 256E / 351Q / 355A, 134V / 353G, 256E, 350S, 353G, and 353M.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and the following sequences compared to SEQ ID NO: 1344: M55R / N58D / C256E / E350S / G373D, N58D, N58S / L69I / C256E / G373D, N58S / D99V / E351Q / I355A , N58S / C256E, E70V / F71A, E70V / F71A / K353G, E70V / S134M / K353G, E70V / K353M, F71A / L77I / K353G, F71A / K353G, L77I, L77I / S134V / K353G, L77I / K353M, D99V / C256E / E351Q / I355A, S134V / K353G, C256E, E350S, K353G, and K353M.

[0149] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and the following sequences compared to SEQ ID NO: 1344: 55 / 58, 55 / 58 / 69 / 350 / 351, 55 / 58 / 99 / 256 / 351 / 373 ... and amino acid sequences having one or more residue differences or sets of residue differences selected from: 6 / 355, 55 / 58 / 99 / 351, 55 / 58 / 256, 55 / 58 / 256 / 355, 55 / 58 / 350 / 351, 58 / 99, 58 / 99 / 355, 58 / 256 / 350 / 355, 58 / 350 / 355, 70, 70 / 353, 71 / 77 / 133 / 353, 71 / 353, 77 / 133 / 353, 77 / 353, 133, 353, and 355 / 373. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and have at least one of the following sequences compared to SEQ ID NO: 1344: 55R / 58M, 55R / 58M / 69I / 350S / 351Q, 55R / 58M / 256E, 55R / 58S / 99V / 256E / 351Q / 373D, 55R / 58S / 99V / 256E / 355A ...M / 69I / 350S / 351Q, 55R / 58M / 256E, 55R / 58S / 99V / 256E / 351Q / 373D, 55R / 58S and 355V / 373D.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and has at least one of the following sequences compared to SEQ ID NO: 1344: M55R / N58M, M55R / N58M / L69I / E350S / E351Q, M55R / N58M / C256E, M55R / N58S / D99V / C256E / E351Q / G373D, M55R / N58S / D99V / C256E / I355A, M55R / N58S / D99V ... 51Q, M55R / N58S / C256E / I355A, M55R / N58S / E350S / E351Q, N58D / D99V / I355V, N58M / C 256E / E350S / I355V, N58S / D99V, N58S / E350S / I355V, E70L, E70V / K353G, F71A / L77I / T and an amino acid sequence having one or more residue differences or a set of residue differences selected from 133Y / K353M, F71A / K353G, L77I / T133Y / K353G, L77I / T133Y / K353M, L77I / K353G, T133Y, K353G, K353M, and I355V / G373D.

[0150] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 1344: 8, 9, 14, 15, 16, 28, 29, 42, 89, 100, 116, 125, 190, 197, 201, 233, 237, 289, 298, 302, 364, 368, and 380. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and the following sequences compared to SEQ ID NO: 1344: 8S, 9D, 14E, 15F, 16F, 17F, 18F, 19F, 20F, 21F, 22F, 23F, 24F, 25F, 26F, 27F, 28F, 29F, 30F, 31F, 32F, 33F, 34F, 35F, 36F, 37F, 38F, 39F, 40F, 41F, 42F, 43F, 44F, 45F, 46F, 47F, 48F, 49F, 50F, 51F, 52F, 53F, 54F, 55F, 56F, 57F, 58F, 59F, 60F, 61F, 62F, 63F, 64F, 65F, 66F, 67F, 68F, 69F, 70F, 71F, 72F, 73F, 74F, 75F, 76F, 77F, 78F, 79F, 80F, 81F, 82F, 83F, 84F, 85F, 86F, 87F, 88F, 89F, , 14G, 15E, 16G, 28G, 29R, 42V, 89G, 89Y, 100E, 116L, 125K, 190V, 197G, 201G, 233R, 237A, 237G, 289G, 298D, 302A, 302G, 364F, 364W, 368S, and 380G. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and the following sequences compared to SEQ ID NO: 1344: G8S, G9D, P14E, P14G, S15E, and an amino acid sequence having one or more residue differences or a set of residue differences selected from N16G, R28G, K29R, N42V, K89G, K89Y, Q100E, R116L, R125K, A190V, N197G, T201G, P233R, D237A, D237G, Y289G, Y298D, K302A, K302G, R364F, R364W, I368S, and A380G.

[0151] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 1344: 9, 14, 28, 86, 89, 91, 92, 104, 116, 125, 196, 197, 198, 201, 230, 234, 248, 266, 272, 278, 298, 304, 307, 308, 341, 364, 365, and 380. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and has at least one of the following sequences compared to SEQ ID NO: 1344: 9E, 14I, 28I, 28S, 86Q, 89D, 91C, 91V, 92A, 92F, 93G, 93H, 93J, 93J, 93K, 93K, 93J ... and amino acid sequences having one or more residue differences or sets of residue differences selected from 2V, 104I, 116T, 125A, 125F, 196C, 196F, 196R, 197R, 198C, 201L, 230D, 230I, 234Q, 248L, 248W, 266Q, 272G, 278S, 298V, 304V, 307L, 308D, 341E, 341T, 364L, 364N, 365G, 380G, and 380M.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1344, and has at least one of the following sequences compared to SEQ ID NO: 1344: G9E, P14I, R28I, R28S, T86Q, K89D, I91C, I91V, Q92A, Q92F, Q92V, V104I ... and amino acid sequences having one or more residue differences or a set of residue differences selected from R116T, R125A, R125F, K196C, K196F, K196R, N197R, T198C, T201L, N230D, N230I, K234Q, K248L, K248W, E266Q, K272G, N278S, Y298V, S304V, A307L, E308D, G341E, G341T, R364L, R364N, K365G, A380G, and A380M.

[0152] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1346 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 1346: 27, 28, 29, 84, 92, 98, 110, 143, 147, 158, 162, 170, 173, 174, 181, 185, 193, 197, 204, 204 / 264 / 340, 290, 297, and 303. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1346, and the following amino acids compared to SEQ ID NO: 1346: 27L, 28E, 28G, 29C, 29P, 84E, 92E, 98A, 99C, 99D, 99E, 99F, 99G, 99H ...100000000000000000000000000000000000000000000000000000000000000000000000 , 98T, 110L, 143A, 147Y, 158R, 162A, 170R, 173L, 174M, 181R, 185R, 193R, 193S, 193V, 197E, 204E / 264L / 340L, 204F, 204I, 204L, 204M, 204R, 290R, 290V, 297A, 303G, and 303T.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1346, and has at least one of the following sequences compared to SEQ ID NO: 1346: A27L, R28E, R28G, K29C, K29P, P84E, Q92E, G98A, G98T, E110 and an amino acid sequence having one or more residue differences or a set of residue differences selected from L, S143A, F147Y, K158R, S162A, K170R, F173L, L174M, S181R, K185R, L193R, L193S, L193V, N197E, P204E / V264L / F340L, P204F, P204I, P204L, P204M, P204R, Q290R, Q290V, S297A, K303G, and K303T.

[0153] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1346 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 1346: 38, 44, 57 / 367, 66, 92, 101, 102, 111, 143, 147, 154, 156, 158, 160, 167, 173, 184, 185, 196, 197, 224, 233, 248, 324, and 352. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1346, and has one of the following sequences compared to SEQ ID NO: 1346: 38R, 44H, 45R ... and amino acid sequences having one or more residue differences or sets of residue differences selected from 57D / 367K, 66G, 92E, 101S, 102K, 111D, 143A, 147Y, 154L, 156D, 158R, 160E, 167R, 173L, 184T, 185R, 196E, 197E, 224V, 233E, 248E, 324I, and 352K. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1346, and has one of the following sequences compared to SEQ ID NO: 1346: K38R, Y44H, E57D / R367K, and an amino acid sequence having one or more residue differences or a set of residue differences selected from I66G, Q92E, V101S, R102K, E111D, S143A, F147Y, V154L, E156D, K158R, S160E, K167R, F173L, S184T, K185R, K196E, N197E, I224V, P233E, K248E, F324I, and R352K.

[0154] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1346 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 1346: 23 / 186 / 256 / 309, 58 / 186 / 270, 182 / 186 / 256 / 360, 182 / 256, 186 / 256, 186 / 256 / 270, 256, and 270 / 309. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1346 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1346: 23V / 186E / 256E / 309K, 58S / 186E / 270T, 182R / 186E / 256E / 360A, 182R / 256E, 186E / 256E, 186E / 256E / 270T, 256E, and 270T / 309K. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1346 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1346: P23V / A186E / C256E / V309K, N58S / A186E / S270T, C182R / A186E / C256E / G360A, C182R / C256E, A186E / C256E, A186E / C256E / S270T, C256E, and S270T / V309K.

[0155] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1678, and one or more residue differences or sets of residue differences compared to SEQ ID NO: 1678 selected from the following: 341. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1678 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1678: 341C, 341H, 341I, 341K, 341L, 341M, 341R, 341T, and 341V. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1678 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1678: G341C, G341H, G341I, G341K, G341L, G341M, G341R, G341T, and G341V.

[0156] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1678, and the following sequences compared to SEQ ID NO: 1678: 9 / 14 / 58, 9 / 14 / 58 / 182 / 190 / 197 / 298, 9 / 14 / 190 / 197 / 36 and 298. The amino acid sequences include those having one or more residue differences or sets of residue differences selected from: 4, 9 / 58 / 197, 9 / 298, 10 / 14 / 197 / 364, 14 / 58 / 182 / 197 / 298 / 364, 14 / 190 / 197 / 298, 14 / 197 / 298 / 364, 14 / 298 / 364, 58 / 197, 58 / 197 / 364, 58 / 364, 182, 182 / 197, 182 / 298 / 364, 197 / 298, and 298. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1678, and the following sequences compared to SEQ ID NO: 1678: 9D / 14I / 58S, 9D / 298V, 9E / 14I / 58S / 182C / 190V / 197G / 298V, 9E / 14I / 190V / 197G / 364L, 9E / and 298V / 364L, 14I / 58S / 182C / 197G / 298V / 364L, 14I / 190V / 197G / 298V, 14I / 197G / 298V / 364L, 14I / 298V / 364F, 58S / 197G, 58S / 197G / 364L, 58S / 364L, 182C, 182C / 197G, 182C / 298V / 364L, 197G / 298V, and 298V.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1678, and has at least one of the following sequences compared to SEQ ID NO: 1678: G9D / P14I / N58S, G9D / Y298V, G9E / P14I / N58S / R182C / A190V / N197G / Y298V, G9E / P14I / A190V / N197G / R364L, G9E / N58S / N197G, and Y298V.

[0157] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1700, and the following sequences compared to SEQ ID NO: 1700: 18 / 28 / 147 / 303, 28, 28 / 84, 28 / 84 / 92 / 147 / 204, 28 / 84 / 156 / 173 / 204 / 290 / 303, 28 / 84 / 173 / 204 / 303, 28 / 84 / 173 / 303, 28 / 84 / 303, 28 / 92 / 147, 28 / 92 / 147 / 204 / 290 / 303, 28 / 92 / 156 / 204 / 290, 28 / 92 / 204, 28 / 147 / 303, 28 / 156 / 204, 28 / 204, 28 / 290, 28 / 303, 29, 29 / 98 / 143 / 266, 29 / 98 / 185 / 197 / 266, 29 / 98 / 185 / 266 / 296 / 299, 29 / 143, 29 / 143 / 162, 29 / 143 / 170 / 193 / 197 / 266, 29 / 143 / 185 / 193 / 266, 29 / 158 / 266, 29 / 170 / 185 / 193 / 266 / 299, 29 / 185 / 296, 29 / 193 / 197 / 296 / 297, 72, 84, 84 / 92 / 173, 84 / 156 / 173 / 204, 84 / 173 / 204 / 303, 92 / 173 / 204 / 290 / 303, 98, 98 / 143 / 158 / 170 / 185 / 296 / 297, 98 / 143 / 162 / 266, 98 / and amino acid sequences having one or more residue differences or sets of residue differences selected from: 143 / 185 / 266, 98 / 162 / 193, 98 / 170 / 193 / 197, 98 / 185, 98 / 185 / 193 / 197 / 266 / 297, 143, 143 / 158 / 197 / 266, 143 / 266 / 296 / 297, 147 / 290 / 303 / 307, 170 / 193 / 197, 173, 173 / 204, 173 / 204 / 303, 182, 204, 266 / 297 / 299, and 303. In some embodiments, the engineered TdT polypeptide has a sequence identity that is at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 110, 111, 112%, 113%, 114%, 115%, 116%, 117%, 118%, 119%, 120%, 121%, 122%, 123%, 124%, 125%, 126%, 127%, 128%, 129%, 130%, 131%, 132%, 133%, 134%, 135%, 136%, 137%, 138%, 139%, 140%, 141%, 142%, 143%, 144%, 145%, 146%, 147%, 148%, 149%, 150%, 151, 152, 153, 154, 155%, 156%, 157%, 158%, 159%, 160%, 161%, 162%, 163%, 164%, 165%, 166%, 167%, 168%, 169%, 170%, 1710%, 1720%, 173%, 174%, 175%, 176%, 177%, 178%, 179%, 180%, 1810%, 1820%,95%, 96%, 97%, 98%, 99% or higher sequence identity, and the following compared to SEQ ID NO: 1700: 18Q / 28G / 147Y / 303G, 28E / 84E, 28E / 84E / 92E / 147Y / 204R, 28E / 84E / 156D / 173L / 204M / 290R / 303G, 28E / 84E / 173L / 303G, 28E / 92E / 147Y, 28E / 290R, 28G, 28G / 84E / 147Y / 204L, 28G / 84E / 173L / 204R / 303T, 28G / 84E / 303G, 28G / 92E / 147Y / 204 I / 290R / 303G, 28G / 92E / 156D / 204L / 290R, 28G / 92E / 204R, 28G / 147Y / 303G , 28G / 156D / 204I, 28G / 204R, 28G / 303G, 29P, 29P / 98A / 185R / 197E / 266G, 29 P / 98T / 143A / 266G, 29P / 98T / 185R / 266G / 296N / 299G, 29P / 143A, 29P / 143A / 162A, 29P / 143A / 170R / 193S / 197E / 266G, 29P / 143A / 185R / 193S / 266G, 29P / 158R / 266G, 29P / 170R / 185R / 193S / 266G / 299G, 29P / 185R / 296N, 29P / 193 V / 197E / 296N / 297A, 72R, 84E, 84E / 92E / 173L, 84E / 156D / 173L / 204F, 84E / 1 73L / 204F / 303T, 92E / 173L / 204F / 290V / 303G, 98A / 185R, 98A / 185R / 193S / 197E / 266G / 297A, 98T, 98T / 143A / 158R / 170R / 185R / 296N / 297A, 98T / 143A / In some embodiments, the amino acid sequence has one or more residue differences or a set of residue differences selected from: 162A / 266G, 98T / 143A / 185R / 266G, 98T / 162A / 193V, 98T / 170R / 193S / 197E, 143A, 143A / 158R / 197E / 266G, 143A / 266G / 296N / 297A, 147Y / 290R / 303T / 307V, 170R / 193S / 197E, 173L, 173L / 204F / 303G, 173L / 204R, 182G, 204R, 266G / 297A / 299G, and 303G.Engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher sequence identity to the reference sequence SEQ ID NO: 1700, and may have any of the following sequences compared to SEQ ID NO: 1700: P18Q / R28G / F147Y / K303G, R28E / P84E, R28E / P84E / Q92E / F147Y / P204R, R28E / P84E / E156D / F173L / P204M / Q290R / K303G, R28E / P84E / F173L / K303G, R28E / Q92E / F147Y, R28E / Q290R, R28G, R28G / P84E / F14 7Y / P204L, R28G / P84E / F173L / P204R / K303T, R28G / P84E / K303G, R28G / Q92E / F147Y / P204I / Q290R / K303G, R28G / Q92E / E156D / P204L / Q290R, R28G / Q92E / P204R, R28G / F147Y / K303G, R28G / E156D / P204I, R28G / P204R, R28G / K303G , K29P, K29P / G98A / K185R / N197E / E266G, K29P / G98T / S143A / E266G, K29P / G 98T / K185R / E266G / S296N / N299G, K29P / S143A, K29P / S143A / S162A, K29P / S 143A / K170R / L193S / N197E / E266G, K29P / S143A / K185R / L193S / E266G, K29P / K158R / E266G, K29P / K170R / K185R / L193S / E266G / N299G, K29P / K185R / S29 6N, K29P / L193V / N197E / S296N / S297A, K72R, P84E, P84E / Q92E / F173L, P84E / E156D / F173L / P204F, P84E / F173L / P204F / K303T, Q92E / F173L / P204F / Q29 0V / K303G, G98A / K185R, G98A / K185R / L193S / N197E / E266G / S297A, G98T, G9 8T / S143A / K158R / K170R / K185R / S296N / S297A, G98T / S143A / S162A / E266G,and amino acid sequences having one or more residue differences or sets of residue differences selected from G98T / S143A / K185R / E266G, G98T / S162A / L193V, G98T / K170R / L193S / N197E, S143A, S143A / K158R / N197E / E266G, S143A / E266G / S296N / S297A, F147Y / Q290R / K303T / A307V, K170R / L193S / N197E, F173L, F173L / P204F / K303G, F173L / P204R, R182G, P204R, E266G / S297A / N299G, and K303G. ,

[0158] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1700, and one or more residue differences or sets of residue differences selected from the following: 29, 60, 72, 84, 147, 182, 211, 220, 224, 266, 290, 338, 339, 342, and 345 relative to SEQ ID NO: 1700. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1700, and has at least one of the following sequences compared to SEQ ID NO: 1700: 29P, 60T, 72E, 72G, 72Q, 72S, 72W, 8 and an amino acid sequence having one or more residue differences or a set of residue differences selected from 4E, 147Y, 182A, 182F, 182G, 182N, 182Q, 182S, 182W, 182Y, 211A, 211V, 220L, 220S, 220V, 220Y, 224A, 224C, 224T, 224V, 266G, 290R, 338A, 338G, 339C, 342A, 345I, and 345K.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1700, and has at least one of the following sequences compared to SEQ ID NO: 1700: K29P, K60T, K72E, K72G, K72Q, K72S, K72W, P84E, F147Y, R and R345K.

[0159] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1750, and the following sequences compared to SEQ ID NO: 1750: 9, 9 / 28 / 156 / 158 / 173 / 190 / 193 / 303 / 364, 9 / 28 / 156 / 158 / 290 / 36 4, 9 / 28 / 190 / 193 / 303 / 364, 9 / 28 / 290 / 303 / 364, 9 / 156 / 158 / 193 / 290 / 303 / 364, 9 / 156 / 364, 9 / 290 / 364, 9 / 364, 14, 14 / 58 / 84 / 147 / 162 / 224 / 296 / 297 / 298 / 299, 14 / 58 / 84 / 147 / 162 / 224 / 296 / 297 / 299, 14 / 58 / 84 / 224 / 298, 14 / 84 / 147 / 193 / 197 / 224 / 296, 14 / 84 / 162 / 224 / 297 / 299, 14 / 84 / 224 / 296 / 299, 14 / 162 / 224 / 298 / 299, 14 / 224 / 296 / 298, 23 / 28 / 156 / 158 / 190 / 193 / 290 / 364, 28 / 156 / 173 / 364, 28 / 158 / 173 / 190 / 193 / 290 / 364, 28 / 190 / 193, 28 / 296 / 303 / 364, 28 / 364, 58 / 147 / 162 / 197 / 224 / 296 / 297 / 2 and amino acid sequences having one or more residue differences or sets of residue differences selected from 98 / 299, 58 / 162 / 224 / 296 / 298, 58 / 224, 58 / 224 / 299, 84 / 147, 147 / 224, 147 / 224 / 297 / 298, 156 / 158 / 190 / 193 / 364, 158 / 193 / 290 / 303, 193 / 290, 224 / 296 / 298, 224 / 297 / 299, 290 / 303 / 364, 303, and 364. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1750, and has the following sequence identity compared to SEQ ID NO: 1750: 9D,9D / 28E / 156D / 158R / 290R / 364L, 9D / 28E / 190V / 193S / 303G / 364L, 9D / 28E / 290R / 303G / 364L, 9D / 28G / 156D / 158R / 173L / 190V / 193S / 303G / 364L, 9D / 156D / 158R / 193V / 290R / 303G / 364L, 9D / 156D / 364L, 9D / 290R / 364L, 9D / 364L, 14I, 14I / 58S / 84E / 147Y / 162A / 224V / 296N / 297A / 298V / 299G, 14I / 58S / 84E / 147Y / 162A / 224V / 296N / 297A / 299G, 14I / 58S / 84E / 224V / 298V, 14I / 84E / 147Y / 193S / 197E / 2 24V / 296N, 14I / 84E / 162A / 224V / 297A / 299G, 14I / 84E / 224V / 296N / 299G, 14I / 162A / 224V / 298V / 299G, 14I / 224V / 296N / 298V, 23S / 2 8E / 156D / 158R / 190V / 193S / 290R / 364L, 28E / 156D / 173L / 364L, 28G / 156D / 173L / 364L, 28G / 158R / 173L / 190V / 193V / 290R / 364L, 28G / 190V / 193V, 28G / 296G / 303G / 364L, 28G / 364L, 58S / 147Y / 162A / 197E / 224V / 296N / 297A / 298V / 299G, 58S / 162A / 224V / 296N / 298V, 5 and amino acid sequences having one or more residue differences or sets of residue differences selected from 8S / 224V, 58S / 224V / 299G, 84E / 147Y, 147Y / 224V, 147Y / 224V / 297A / 298V, 156D / 158R / 190V / 193V / 364L, 158R / 193V / 290R / 303G, 193V / 290R, 224V / 296N / 298V, 224V / 297A / 299G, 290R / 303G / 364L, 303T, and 364L. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1750.and the following relative to SEQ ID NO: 1750: G9D, G9D / R28E / E156D / K158R / Q290R / R364L, G9D / R28E / A190V / L193S / K303G / R364L, G9D / R28E / Q290R / K303G / R364L, G9D / R28G / E156D / K158R / F173L / A190V / L193S / K303G / R364L, G9D / E156D / K158R / L193V / Q290R / K303G / R364L, G9D / E156D / K158R / L193V / Q290R / K303G / R364L, G9D / Q290R / R364L, G9D / R36 4L, P14I, P14I / N58S / P84E / F147Y / S162A / I224V / S296N / S297A / Y298V / N2 99G, P14I / N58S / P84E / F147Y / S162A / I224V / S296N / S297A / N299G, P14I / N5 8S / P84E / I224V / Y298V, P14I / P84E / F147Y / L193S / N197E / I224V / S296N, P1 4I / P84E / S162A / I224V / S297A / N299G, P14I / P84E / I224V / S296N / N299G, P1 4I / S162A / I224V / Y298V / N299G, P14I / I224V / S296N / Y298V, P23S / R28E / E 156D / K158R / A190V / L193S / Q290R / R364L, R28E / E156D / F173L / R364L, R28G / E156D / F173L / R364L, R28G / K158R / F173L / A190V / L193V / Q290R / R364L, R 28G / A190V / L193V, R28G / S296G / K303G / R364L, R28G / R364L, N58S / F147Y / S 162A / N197E / I224V / S296N / S297A / Y298V / N299G, N58S / S162A / I224V / S29 6N / Y298V, N58S / I224V, N58S / I224V / N299G, P84E / F147Y, F147Y / I224V, F1 47Y / I224V / S297A / Y298V, E156D / K158R / A190V / L193V / R364L, K158R / L193 V / Q290R / K303G, L193V / Q290R, I224V / S296N / Y298V, I224V / S297A / N299G,The amino acid sequence has one or more residue differences or a set of residue differences selected from Q290R / K303G / R364L, K303T, and R364L.

[0160] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1750, and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 1750: 34, 48, 69, 78, 237 / 271, 249, 302, 309, 315, 353, 364, and 365. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1750 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1750: 34A, 48L, 69I, 78F, 237A / 271H, 249T, 302G, 309K, 315V, 353Q, 364F, 364L, and 365G. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1750, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1750: S34A, F48L, L69I, L78F, D237A / R271H, K249T, K302G, V309K, I315V, G353Q, R364F, R364L, and K365G.

[0161] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1932, and has the following sequence identity compared to SEQ ID NO: 1932: 12, 14, 14 / 28 / 58 / 84 / 173 / 297, 14 / 28 / 58 / 162 / 190 / 193 / 224, 14 / 28 / 58 / 224, 14 / 28 / 224 / 296 / 298, 14 / 58 / 147 / 162 / 190 / 193 / 224 / 296 / 303, 14 / 58 / 158 / 224, 14 / 58 / 162, 14 / 147, 14 / 147 / 158 / 162 / 224, 14 / 147 / 173, 14 / 147 / 224, 14 / 158 / 162 / 197 / 224 / 296 / 298 / 299, 14 / 173 / 193 / 197 / 296 / 297 / 298 / 299, 14 / 193 / 197 / 224 / 297 / 298 / 299 / 300, 14 / 224 / 296 / 303, 28, 28 / 58 / 84 / 158 / 162 / 193 / 224 / 296 / 298 / 299, 28 / 58 / 147 / 224 / 296 / 297 / 298 / 299 / 303, 28 / 58 / 162 / 224 / 298 / 299, 28 / 84, 28 / 84 / 147 / 297 / 298, 28 / 224 / 296 / 297 / 298 / 303, 28 / 296 / 298, 58 / 72 / 211 / 315, 58 / 72 / 220 / 224, 58 / 84 / 211 / 220 / 224, 58 / 147 / 162 / 296 / 298, 66 / 220 / 224, 72, 72 / 84 / 86 / 224, 72 / 84 / 220 / 224 / 315, 72 / 86 / 220, 72 / 220 / 224, 72 / 220 / 315, 75, 77, 84 / 147 / 197 / 296 / 297, 84 / 147 / 297 / 298 / 303, 84 / 173 / 224, 84 / 220 / 315, 84 / 224 / 297 / 298 / 299, 100, 104, 120, 147, 147 / 158 / 162 / 1 90 / 224 / 296 / 297 / 299 / 303, 162 / 224, 173, 190 / 193 / 197, 193 / 197 / 296 / 303, 197, 220 / 224 / 315, 220 / 315, 224, 224 / 298 / 299, 233, 292, 297, 315, 353,and 367. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1932, and one or more of the following residue differences or sets of residue differences compared to SEQ ID NO: 1932: 12L, 14I, 14I / 28G / 58S / 84E / 173L / 297A, 14I / 28G / 58S / 162A / 190V / 193V / 224V, 14I / 28G / 58S ... 14I / 28G / 224V / 296N / 298V, 14I / 58S / 147Y / 162A / 190V / 193V / 224V / 296 N / 303G, 14I / 58S / 158R / 224V, 14I / 58S / 162A, 14I / 147Y, 14I / 147Y / 158 R / 162A / 224V, 14I / 147Y / 173L, 14I / 147Y / 224V, 14I / 158R / 162A / 197E / 224V / 296N / 298V / 299G, 14I / 173L / 193V / 197E / 296N / 297A / 298V / 299G, 14I / 193V / 197E / 224V / 297R / 298W / 299~ / 300A, 14I / 224V / 296N / 303G, 2 8E / 58S / 84E / 158R / 162A / 193V / 224V / 296N / 298V / 299G, 28E / 58S / 147Y / 224V / 296N / 297A / 298V / 299G / 303G, 28E / 58S / 162A / 224V / 298V / 299G, 2 8G, 28G / 84E, 28G / 84E / 147Y / 297A / 298V, 28G / 224V / 296N / 297A / 298V / 3 03G, 28G / 296N / 298V, 58D / 72Q / 211A / 315V, 58D / 72Q / 220V / 224V, 58D / 8 4E / 211A / 220V / 224V, 58S / 147Y / 162A / 296N / 298V, 66P / 220V / 224V, 72G / 84E / 86Q / 224V, 72G / 84E / 220V / 224V / 315V, 72Q, 72Q / 86Q / 220V, 72Q / 2 20V / 224V, 72Q / 220V / 315V, 75V, 77L, 77V, 84E / 147Y / 197E / 296N / 297A,84E / 147Y / 297A / 298V / 303G, 84E / 173L / 224V, 84E / 220V / 315V, 84E / 224V / 297A / 298V / 299G, 100D , 104L, 120I, 147Y, 147Y / 158R / 162A / 190V / 224V / 296N / 297A / 299G / 303G, 162A / 224V, 173L, 190V / 193V / 197E, 193V / 197E / 296N / 303G, 197E, 220V / 224V / 315V, 220V / 315V, 224V, 224V / 298V / 299G, 233G, 233R, 292K, 297A, 315V, 353D, and 367K. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1932, and has at least one of the following sequences compared to SEQ ID NO: 1932: H12L, P14I, P14I / R28G / N58S / P84E / F173L / S297A, P14I / R28G / N58S / S162A / A190V / L193V / I224V, P14I / R28G / N58S / I 224V, P14I / R28G / I224V / S296N / Y298V, P14I / N58S / F147Y / S162A / A190V / L193V / I224V / S296N / K303G , P14I / N58S / K158R / I224V, P14I / N58S / S162A, P14I / F147Y, P14I / F147Y / K158R / S162A / I224V, P14I / F147Y / F173L, P14I / F147Y / I224V, P14I / K158R / S162A / N197E / I224V / S296N / Y298V / N299G, P14I / F1 73L / L193V / N197E / S296N / S297A / Y298V / N299G, P14I / L193V / N197E / I224V / S297R / Y298W / N299~ / T3 00A, P14I / I224V / S296N / K303G, R28E / N58S / P84E / K158R / S162A / L193V / I224V / S296N / Y298V / N299G,R28E / N58S / F147Y / I224V / S296N / S297A / Y298V / N299G / K303G, R28E / N58S / S162A / I224V / Y298V / N299G, R28G, R28G / P84E, R2 8G / P84E / F147Y / S297A / Y298V, R28G / I224V / S296N / S297A / Y298V / K303G, R28G / S296N / Y298V, N58D / K72Q / T211A / I315V, N58 D / K72Q / M220V / I224V, N58D / P84E / T211A / M220V / I224V, N58S / F147Y / S162A / S296N / Y298V, I66P / M220V / I224V, K72G / P84E / T86Q / I224V, K72G / P84E / M220V / I224V / I315V, K72Q, K72Q / T86Q / M220V, K72Q / M220V / I224V, K72Q / M220V / I315V, A75V, I77L , I77V, P84E / F147Y / N197E / S296N / S297A, P84E / F147Y / S297A / Y298V / K303G, P84E / F173L / I224V, P84E / M220V / I315V, P84E / I224V / S297A / Y298V / N299G, Q100D, V104L, V120I, F147Y, F147Y / K158R / S162A / A190V / I224V / S296N / S297A / N299G / K303G, S and an amino acid sequence having one or more residue differences or a set of residue differences selected from: 162A / I224V, F173L, A190V / L193V / N197E, L193V / N197E / S296N / K303G, N197E, M220V / I224V / I315V, M220V / I315V, I224V, I224V / Y298V / N299G, P233G, P233R, G292K, S297A, I315V, G353D, and R367K.

[0162] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1932 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 1932: 12, 75, 77, 100, 104, 120, 197, 233, 292, 353, and 367. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1932 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1932: 12L, 75V, 77L, 77V, 100D, 104L, 120I, 197E, 233G, 233R, 292K, 353D, and 367K. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 1932 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 1932: H12L, A75V, I77L, I77V, Q100D, V104L, V120I, N197E, P233G, P233R, G292K, G353D, and R367K.

[0163] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:2164, and one or more residue differences or sets of residue differences selected from the following: 51, 52, 66, and 71 compared to SEQ ID NO:2164. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: 51V, 52V, 66E, 66S, and 71M. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: A51V, L52V, I66E, I66S, and A71M.

[0164] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164, and the following: 9 / 11, 20, 52, 74, 78, 80 , 96, 101, 108, 111, 116, 145, 147, 173, 200, 203, 206, 232, 235, 242, 249, 264, 267, 288, 293, 304, 306, 308, 324, 326, 328, 350, 352, 361, 368, 374, 378, and 380. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164, and has at least one of the following sequences compared to SEQ ID NO: 2164: 9D / 11L, 20E, 20K, 20L, 20R, 20V, 52G, 74L, 78M, 80S, 96A, 101C, 108D, 111F, 111R, 116L, 116P, 145C, 146D, 147D, 148D, 148F, 148H ... and 380S.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164, and has at least one of the following sequences compared to SEQ ID NO: 2164: G9D / G11L, G20E, G20K, G20L, G20R, G20V, L52G, A74L, L78M, Y80S, C96A, V101C, I108D, E111F, E111R, R116L, R116P, K145C, K145R, K14 and A380S, and the amino acid sequence has one or more residue differences or a set of residue differences selected from: 5V, F147M, F173R, R200I, R200L, L203S, L203V, A206V, G232M, E235C, E235F, E235M, E235R, E235V, K242R, K249A, K249G, V264T, Q267S, L288R, V293D, V293G, S304G, S304L, M306P, E308L, F324V, Q326C, A328W, E350L, E350T, R352A, L361W, I368R, S374D, I378G, and A380S.

[0165] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:2164, and one or more residue differences or sets of residue differences selected from the following: 97, 168, 259, 273, and 276 compared to SEQ ID NO:2164. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: 97L, 168A, 168Q, 259Q, 259V, 273V, and 276L. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: I97L, M168A, M168Q, K259Q, K259V, I273V, and M276L.

[0166] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:2164, and one or more residue differences or sets of residue differences selected from the following: 97, 135, 259, 273, 276, and 278 compared to SEQ ID NO:2164. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: 97L, 135I, 259Q, 259V, 273V, 276L, and 278A. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: I97L, V135I, K259Q, K259V, I273V, M276L, and N278A.

[0167] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:2164, and one or more residue differences or sets of residue differences selected from the following: 97, 135, 166, 273, and 278 compared to SEQ ID NO:2164. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: 97L, 135I, 166T, 273V, and 278A. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: I97L, V135I, S166T, I273V, and N278A.

[0168] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:2164 and one or more residue differences or sets of residue differences selected from the following: 97, 135, 140, 166, 167, 168, 230, 259, 273, 276, and 278 compared to SEQ ID NO:2164. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: 97F, 97L, 135I, 140L, 166N, 167R, 168Q, 230H, 259Q, 259V, 273V, 276L, 278A, and 278R. In some embodiments, an engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2164: I97F, I97L, V135I, V140L, S166N, K167R, M168Q, N230H, K259Q, K259V, I273V, M276L, N278A, and N278R.

[0169] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164, and has at least one of the following sequences compared to SEQ ID NO: 2164: 34, 34 / 48 / 78 / 133 / 147 / 182, 34 / 48 / 78 / 147 / 158, 34 / 48 / 78 / 147 / 182, 220, 233, 249, 307, 315, 339, 34 / 48 / 78 / 158 / 182 / 220 / 249 / 307, 34 / 48 / 78 / 158 / 182 / 233 / 315 / 345, 34 / 48 / 78 / 182 / 220 / 230 / 315, 34 / 48 / 78 / 182 / 220 / 233, 34 / 48 / 133 / 158 / 182 / 230 / 233 / 271 / 34 5, 34 / 48 / 147, 34 / 48 / 147 / 158 / 182 / 220 / 233 / 307, 34 / 48 / 147 / 182 / 230 / 233 / 249 / 307, 34 / 48 / 147 / 271, 34 / 48 / 182 / 233 / 249, 34 / 48 / 339, 34 / 78, 34 / 7 8 / 84 / 147 / 158 / 182 / 220, 34 / 78 / 84 / 158 / 230, 34 / 78 / 147 / 182 / 220 / 249, 34 / 78 / 147 / 182 / 233 / 249 / 315, 34 / 78 / 147 / 182 / 233 / 271 / 339, 34 / 78 / 158, 3 4 / 78 / 158 / 182 / 315 / 345, 34 / 78 / 158 / 249, 34 / 78 / 182 / 233 / 307, 34 / 78 / 204 / 220 / 339, 34 / 78 / 220 / 271, 34 / 78 / 220 / 307 / 339, 34 / 78 / 315, 34 / 133 / 147 / 158 / 230 / 233 / 249, 34 / 147, 34 / 147 / 155 / 233 / 339, 34 / 147 / 158, 34 / 147 / 158 / 182 / 233, 34 / 147 / 158 / 182 / 249 / 271, 34 / 147 / 182, 34 / 147 / 182 / 220, 34 / 147 / 182 / 220 / 230 / 249 / 315 / 339, 34 / 147 / 182 / 220 / 230 / 339, 34 / 147 / 182 / 220 / 271 / 315, 34 / 147 / 182 / 233 / 271 / 339, 34 / 147 / 182 / 249 / 307 / 315 / 339,34 / 147 / 182 / 315、34 / 147 / 182 / 345、34 / 147 / 220 / 271、34 / 147 / 230 / 273 / 315 / 345、34 / 147 / 233、34 / 147 / 249、34 / 147 / 315 / 339、34 / 158 / 182 / 315 / 339 / 345、34 / 158 / 220、34 / 158 / 307、34 / 182、34 / 182 / 230 / 315、34 / 182 / 307 / 339、34 / 182 / 345、34 / 220、34 / 220 / 307、34 / 220 / 307 / 345、34 / 220 / 315 / 339、34 / 220 / 339、34 / 233、34 / 271 / 339、34 / 315 / 345、34 / 339、48 / 78 / 147 / 158 / 182 / 220 / 230 / 307、48 / 78 / 147 / 158 / 182 / 230 / 249 / 271 / 315、48 / 78 / 147 / 158 / 233 / 249、48 / 78 / 147 / 182 / 220 / 233 / 249 / 339 / 345、48 / 78 / 147 / 182 / 220 / 315、48 / 78 / 147 / 182 / 230 / 233 / 249、48 / 78 / 158 / 182 / 220 / 233 / 249、48 / 78 / 158 / 230 / 339、48 / 78 / 233 / 315 / 339、48 / 147 / 158 / 182 / 220 / 230 / 249 / 271 / 307 / 315 / 339、48 / 147 / 158 / 182 / 220 / 249 / 307 / 339、48 / 147 / 158 / 182 / 220 / 315、48 / 147 / 158 / 182 / 230 / 233、48 / 147 / 158 / 182 / 233 / 345、48 / 147 / 158 / 233、48 / 147 / 158 / 307 / 345、48 / 147 / 233、48 / 147 / 233 / 345、48 / 147 / 271 / 307 / 339、48 / 158 / 182 / 230 / 233 / 249、48 / 182、48 / 182 / 307 / 315、48 / 339、78 / 133 / 147 / 158 / 182 / 220 / 271 / 339、78 / 135 / 182 / 233 / 249 / 315 / 345、78 / 147 / 158 / 182、78 / 147 / 158 / 182 / 230 / 249、78 / 147 / 158 / 182 / 233 / 271 / 307 / 345、78 / 147 / 158 / 182 / 339 / 345、78 / 147 / 158 / 220 / 230 / 233 / 249 / 271 / 307 / 315 / 345、78 / 147 / 158 / 249 / 307 / 315, 78 / 147 / 182 / 230 / 233 / 249, 78 / 147 / 182 / 249 / 307, 78 / 147 / 182 / 339 / 345, 78 / 147 / 230 / 307, 78 / 147 / 233, 78 / 147 / 249 / 271 / 339, 78 / 158 / 182, 78 / 158 / 182 / 233 / 271 / 315, 78 / 158 / 182 / 307 / 315 / 345, 78 / 182 / 220 / 339, 78 / 182 / 271 / 315 / 339, 78 / 233, 78 / 339, 133 / 135 / 307 / 315, 133 / 220 / 233 / 271, 133 / 307, 147, 147 / 158 / 182 / 220 / 233, 147 / 158 / 182 / 230 / 233 / 249 / 271 / 339, 147 / 158 / 182 / 233 / 249, 147 / 158 / 182 / 233 / 271 / 307 / 339, 147 / 158 / 182 / 233 / 307 / 339, 147 / 158 / 182 / 233 / 315 / 345, 147 / 158 / 182 / 271 / 315, 147 / 158 / 182 / 3 15, 147 / 158 / 220 / 230 / 233 / 249 / 345, 147 / 158 / 220 / 249 / 315, 147 / 158 / 233 / 249, 147 / 182 / 220 / 233 / 345, 147 / 182 / 230 / 307 / 315 / 339, 147 / 182 / 233, 147 / 182 / 233 / 249, 147 / 182 / 249 / 271 / 307, 147 / 182 / 345, 147 / 220, 147 / 232 / 233 / 271 / 315 / 339, 147 / 233 / 345, 147 / 249, 147 / 339 , 158 / 182 / 220 / 230 / 249 / 307 / 339, 158 / 233, 158 / 307, 158 / 315, 158 / 315 / 339, 182, 182 / 220 / 345, 182 / 230, 182 / 230 / 233, 182 / 345, 220, 220 / 249, 220 / 307 / 339, 233, 233 / 271, 233 / 315, 249, 249 / 315 / 339, 271 / 339, 315, and 339. In some embodiments, the engineered TdT polypeptide has a sequence identity that is at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher sequence identity, and the following compared to SEQ ID NO: 2164: 34A, 34A / 48L / 78F / 133S / 147Y / 182M, 34A / 48L / 78F / 147Y / 158R, 34A / 48L / 78F / 147Y / 182M / 220V / 233G / 249T / 307G / 315V / 339Q, 34A / 48L / 78F / 158R / 182G / 233R / 315V / 345K, 34A / 48L / 78F / 158R / 182M / 220V / 249T / 307 G, 34A / 48L / 78F / 182G / 220V / 230L / 315V, 34A / 48L / 78F / 182G / 220V / 233R, 3 4A / 48L / 133S / 158R / 182G / 230L / 233G / 271H / 345K, 34A / 48L / 147Y, 34A / 48L / 147Y / 158R / 182M / 220V / 233R / 307G, 34A / 48L / 147Y / 182G / 230L / 233G / 249 T / 307G, 34A / 48L / 147Y / 271H, 34A / 48L / 182G / 233G / 249T, 34A / 48L / 339Q, 3 4A / 78F, 34A / 78F / 84G / 147Y / 158R / 182M / 220V, 34A / 78F / 84G / 158R / 230L, 3 4A / 78F / 147Y / 182G / 233R / 249T / 315V, 34A / 78F / 147Y / 182M / 220V / 249T, 34 A / 78F / 147Y / 182M / 233R / 271H / 339Q, 34A / 78F / 158R, 34A / 78F / 158R / 182M / 315V / 345K, 34A / 78F / 158R / 249T, 34A / 78F / 182M / 233R / 307G, 34A / 78F / 204 T / 220V / 339Q, 34A / 78F / 220V / 271H, 34A / 78F / 220V / 307G / 339Q, 34A / 78F / 3 15V, 34A / 133S / 147Y / 158R / 230L / 233G / 249T, 34A / 147Y, 34A / 147Y / 155D / 2 33G / 339Q, 34A / 147Y / 158R, 34A / 147Y / 158R / 182G / 249T / 271H, 34A / 147Y / 1 58R / 182M / 233R, 34A / 147Y / 182G / 220V, 34A / 147Y / 182G / 233G / 271H / 339Q,34A / 147Y / 182G / 345K、34A / 147Y / 182M、34A / 147Y / 182M / 220V / 230L / 249T / 315V / 339Q、34A / 147Y / 182M / 220V / 230L / 339Q、34A / 147Y / 182M / 220V / 271H / 315V、34A / 147Y / 182M / 249T / 307G / 315V / 339Q、34A / 147Y / 182M / 315V、34A / 147Y / 220V / 271H、34A / 147Y / 230L / 273N / 315V / 345K、34A / 147Y / 233R、34A / 147Y / 249T、34A / 147Y / 315V / 339Q、34A / 158R / 182G / 315V / 339Q / 345K、34A / 158R / 220V、34A / 158R / 307G、34A / 182G / 307G / 339Q、34A / 182G / 345K、34A / 182M、34A / 182M / 230L / 315V、34A / 220V、34A / 220V / 307G、34A / 220V / 307G / 345K、34A / 220V / 315V / 339Q、34A / 220V / 339Q、34A / 233G、34A / 271H / 339Q、34A / 315V / 345K、34A / 339Q、48L / 78F / 147Y / 158M / 182G / 230L / 249T / 271H / 315V、48L / 78F / 147Y / 158R / 182M / 220V / 230L / 307G、48L / 78F / 147Y / 158R / 233G / 249T、48L / 78F / 147Y / 182G / 230L / 233G / 249T、48L / 78F / 147Y / 182M / 220V / 233G / 249T / 339Q / 345K、48L / 78F / 147Y / 182M / 220V / 315V、48L / 78F / 158R / 182G / 220V / 233G / 249T、48L / 78F / 158R / 230L / 339Q、48L / 78F / 233R / 315V / 339Q、48L / 147Y / 158R / 182G / 220V / 249T / 307G / 339Q、48L / 147Y / 158R / 182G / 230L / 233G、48L / 147Y / 158R / 182M / 220V / 230L / 249T / 271H / 307G / 315V / 339Q、48L / 147Y / 158R / 182M / 220V / 315V、48L / 147Y / 158R / 182M / 233G / 345K、48L / 147Y / 158R / 233G、48L / 147Y / 158R / 307G / 345K、48L / 147Y / 233G / 345K、48L / 147Y / 233R、48L / 147Y / 271H / 307G / 339Q、48L / 158R / 182M / 230L / 233R / 249T、48L / 182G / 307G / 315V、48L / 182M、4、 8L / 339Q、78F / 133S / 147Y / 158R / 182G / 220V / 271H / 339Q、78F / 135A / 182M / 233R / 249T / 315V / 345K、78F / 147Y / 158R / 182G / 230L / 249T、78F / 147Y / 158R / 182G / 339Q / 345K、78F / 147Y / 158R / 182M、78F / 147Y / 158R / 182M / 233R / 271H / 307G / 345K、78F / 147Y / 158R / 220V / 230L / 233G / 249T / 271H / 307G / 315V / 345K、78F / 147Y / 158R / 249T / 307G / 315V、78F / 147Y / 182G / 249T / 307G、78F / 147Y / 182M / 230L / 233G / 249T、78F / 147Y / 182M / 339Q / 345K、78F / 147Y / 230L / 307G、78F / 147Y / 233G、78F / 147Y / 249T / 271H / 339Q、78F / 158R / 182M、78F / 158R / 182M / 233R / 271H / 315V、78F / 158R / 182M / 307G / 315V / 345K、78F / 182G / 271H / 315V / 339Q、78F / 182M / 220V / 339Q、78F / 233R、78F / 339Q、133S / 135A / 307G / 315V、133S / 220V / 233G / 271H、133S / 307G、147Y、147Y / 158R / 182G / 220V / 233G、147Y / 158R / 182G / 233G / 271H / 307G / 339Q、147Y / 158R / 182G / 233G / 307G / 339Q、147Y / 158R / 182M / 230L / 233R / 249T / 271H / 339Q、147Y / 158R / 182M / 233R / 249T、147Y / 158R / 182M / 233R / 315V / 345K、147Y / 158R / 182M / 271H / 315V、147Y / 158R / 182M / 315V、147Y / 158R / 220V / 230L / 233R / 249T / 345K、147Y / 158R / 220V / 249T / 315V、147Y / 158R / 233G / 249T、147Y / 182G / 233R、147Y / 182M / 220V / 233R / 345K、147Y / 182M / 230L / 307G / 315V / 339Q、147Y / 182M / 233G / 249T, 147Y / 182M / 249T / 271H / 307G, 147Y / 182M / 345K, 147Y / 220V, 147Y / 232N / 233G / 271H / 315V / 339Q, 14 7Y / 233R / 345K, 147Y / 249T, 147Y / 339Q, 158R / 182G / 220V / 230L / 249T / 307G / 339Q, 158R / 233R, 158R / 307G, 158R / 315V, 158R / and amino acid sequences having one or more residue differences or sets of residue differences selected from 315V / 339Q, 182G, 182G / 230L / 233R, 182G / 345K, 182M / 220V / 345K, 182M / 230L, 220V, 220V / 249T, 220V / 307G / 339Q, 233G / 315V, 233R, 233R / 271H, 249T, 249T / 315V / 339Q, 271H / 339Q, 315V, and 339Q. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2164, and has at least one of the following sequences compared to SEQ ID NO: 2164: S34A, S34A / F48L / L78F / T133S / F147Y / R182M, S34A / F48L / L78F / F147Y / K158R, S34A / F48L / L78F / F147Y / R182M / M220V / P233G / K249T / A307G / I315V / E33 9Q, S34A / F48L / L78F / K158R / R182G / P233R / I315V / R345K, S34A / F48L / L78F / K158R / R182M / M220V / K249T / A307G, S34A / F48L / L78F / R182G / M220V / N230L / I315V, S34A / F4 8L / L78F / R182G / M220V / P233R, S34A / F48L / T133S / K158R / R182G / N230L / P233G / R271 H / R345K, S34A / F48L / F147Y, S34A / F48L / F147Y / K158R / R182M / M220V / P233R / A307G,S34A / F48L / F147Y / R182G / N230L / P233G / K249T / A307G、S34A / F48L / F147Y / R271H、S34A / F48L / R182G / P233G / K249T、S34A / F48L / E339Q、S34A / L78F、S3 4A / L78F / E84G / F147Y / K158R / R182M / M220V、S34A / L78F / E84G / K158R / N230 L、S34A / L78F / F147Y / R182G / P233R / K249T / I315V、S34A / L78F / F147Y / R182 M / M220V / K249T、S34A / L78F / F147Y / R182M / P233R / R271H / E339Q、S34A / L78 F / K158R、S34A / L78F / K158R / R182M / I315V / R345K、S34A / L78F / K158R / K249 T、S34A / L78F / R182M / P233R / A307G、S34A / L78F / P204T / M220V / E339Q、S34A / L78F / M220V / R271H、S34A / L78F / M220V / A307G / E339Q、S34A / L78F / I315V、S 34A / T133S / F147Y / K158R / N230L / P233G / K249T、S34A / F147Y、S34A / F147Y / E155D / P233G / E339Q、S34A / F147Y / K158R、S34A / F147Y / K158R / R182G / K249 T / R271H、S34A / F147Y / K158R / R182M / P233R、S34A / F147Y / R182G / M220V、S3 4A / F147Y / R182G / P233G / R271H / E339Q、S34A / F147Y / R182G / R345K、S34A / F1 47Y / R182M、S34A / F147Y / R182M / M220V / N230L / K249T / I315V / E339Q、S34A / F147Y / R182M / M220V / N230L / E339Q、S34A / F147Y / R182M / M220V / R271H / I31 5V、S34A / F147Y / R182M / K249T / A307G / I315V / E339Q、S34A / F147Y / R182M / I315V、S34A / F147Y / M220V / R271H、S34A / F147Y / N230L / I273N / I315V / R345K、S34A / F147Y / P233R、S34A / F147Y / K249T、S34A / F147Y / I315V / E339Q、S34A / K158R / R182G / I315V / E339Q / R345K、S34A / K158R / M220V、S34A / K158R / A307G、S34A / R182G / A307G / E339Q、S34A / R182G / R345K、S34A / R182M、S34A / R182M / N230L / I315V、S34A / M220V、S34A / M220V / A307G、S34A / M220V / A307G / R345K、S34A / M220V / I315V / E339Q、S34A / M220V / E339Q、S34A / P233G、S34A / R271H / E339Q、S34A / I315V / R345K、S34A / E339Q、F48L / L78F / F147Y / K158M / R182G / N230L / K249T / R271H / I315V、F48L / L78F / F147Y / K158R / R182M / M220V / N230L / A307G、F48L / L78F / F147Y / K158R / P233G / K249T、F48L / L78F / F147Y / R182G / N230L / P233G / K249T、F48L / L78F / F147Y / R182M / M220V / P233G / K249T / E339Q / R345K、F48L / L78F / F147Y / R182M / M220V / I315V、F48L / L78F / K158R / R182G / M220V / P233G / K249T、F48L / L78F / K158R / N230L / E339Q、F48L / L78F / P233R / I315V / E339Q、F48L / F147Y / K158R / R182G / M220V / K249T / A307G / E339Q、F48L / F147Y / K158R / R182G / N230L / P233G、F48L / F147Y / K158R / R182M / M220V / N230L / K249T / R271H / A307G / I315V / E339Q、F48L / F147Y / K158R / R182M / M220V / I315V、F48L / F147Y / K158R / R182M / P233G / R345K、F48L / F147Y / K158R / P233G、F48L / F147Y / K158R / A307G / R345K、F48L / F147Y / P233G / R345K、F48L / F147Y / P233R、F48L / F147Y / R271H / A307G / E339Q、F48L / K158R / R182M / N230L / P233R / K249T、F48L / R182G / A307G / I315V、F48L / R182M、F48L / E339Q、L78F / T133S / F147Y / K158R / R182G / M220V / R271H / E339Q、L78F / V135A / R182M / P233R / K249T / I315V / R345K、L78F / F147Y / K158R / R182G / N230L / K249T、L78F / F147Y / K158R / R182G / E339Q / R345K、L78F / F147Y / K158R / R182M、L78F / F147Y / K158R / R182M / P233R / R271H / A307G / R345K、L78F / F147Y / K158R / M220V / N230L / P233G / K249T / R271H / A307G / I315V / R345K、L78F / F147Y / K158R / K249T / A307G / I315V、L78F / F147Y / R182G / K249T / A307G、L78F / F147Y / R182M / N230L / P233G / K249T、L78F / F147Y / R182M / E339Q / R345K、L78F / F147Y / N230L / A307G、L78F / F147Y / P233G、L78F / F147Y / K249T / R271H / E339Q、L78F / K158R / R182M、L78F / K158R / R182M / P233R / R271H / I315V、L78F / K158R / R182M / A307G / I315V / R345K、L78F / R182G / R271H / I315V / E339Q、L78F / R182M / M220V / E339Q、L78F / P233R、L78F / E339Q、T133S / V135A / A307G / I315V、T133S / M220V / P233G / R271H、T133S / A307G、F147Y、F147Y / K158R / R182G / M220V / P233G、F147Y / K158R / R182G / P233G / R271H / A307G / E339Q、F147Y / K158R / R182G / P233G / A307G / E339Q、F147Y / K158R / R182M / N230L / P233R / K249T / R271H / E339Q、F147Y / K158R / R182M / P233R / K249T, F147Y / K158R / R182M / P233R / I315V / R345K, F147Y / K158R / R182M / R271H / I31 5V, F147Y / K158R / R182M / I315V, F147Y / K158R / M220V / N230L / P233R / K249T / R345K, F147Y / K158R / M220V / K249T, / I315V, F147Y / K158R / P233G / K249T, F147Y / R182G / P233R, F147Y / R182M / M220V / P233R / R 345K, F147Y / R182M / N230L / A307G / I315V / E339Q, F147Y / R182M / P233G / K249T, F147Y / R182 M / K249T / R271H / A307G, F147Y / R182M / R345K, F147Y / M220V, F147Y / G232N / P233G / R271H / I 315V / E339Q, F147Y / P233R / R345K, F147Y / K249T, F147Y / E339Q, K158R / R182G / M220V / N230 L / K249T / A307G / E339Q, K158R / P233R, K158R / A307G, K158R / I315V, K158R / I315V / E339Q, R182G, R182G / N230L / P233R, R182G / R345K, R182M / M220V / R345K, R182M / N230L, M220V, M22 and an amino acid sequence having one or more residue differences or a set of residue differences selected from: 0V / K249T, M220V / A307G / E339Q, P233G / I315V, P233R, P233R / R271H, K249T, K249T / I315V / E339Q, R271H / E339Q, I315V, and E339Q.

[0170] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 2666: 162, 167, 259, 272, 273, 345, 346, and 365. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2666: 162T, 167A, 259R, 272M, 273E, 345C, 346W, and 365G. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2666: S162T, K167A, K259R, K272M, I273E, R345C, R346W, and K365G.

[0171] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or sets of residue differences selected from the following: 260, 272, 273, 280, 345, and 346 compared to SEQ ID NO: 2666. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2666: 260P, 272M, 273E, 280S, 345C, 346Q, and 346W. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2666: E260P, K272M, I273E, Q280S, R345C, R346Q, and R346W.

[0172] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:2666, and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO:2666: 167, 170, 259, 260, 272, 325, 346, 365, 367, 368, 371, and 373. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2666: 167R, 170R, 259R, 260P, 272G, 272M, 325S, 346Q, 365S, 367G, 368S, 371P, and 373M. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2666: K167R, K170R, K259R, E260P, K272G, K272M, E325S, R346Q, K365S, R367G, I368S, K371P, and G373M.

[0173] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 2666: 162, 259, 272, 273, 325, 345, 346, 367, and 373. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2666: 162T, 259R, 272G, 272M, 273E, 325S, 345C, 346Q, 346W, 367G, and 373M. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2666: S162T, K259R, K272G, K272M, I273E, E325S, R345C, R346Q, R346W, R367G, and G373M.

[0174] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:2666, and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO:2666: 136, 162, 164, 167, 170, 200 / 271, 259, 262, 267, 268, 272, 278, 284, 321, 324, 327, 328, 345, 346, 352, 353, 355, 364, 365, 366, 368, 370, 371, and 373. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and has the following sequences compared to SEQ ID NO: 2666: 136W, 162C, 162E, 162F, 162H, 162T, 162V, 164A, 164S, 164W, 167A, 170T, 200C / 200D, 200E / 200F, 200H / 200G, 200H / 200H, 200I / 200I ... and 373S.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and has at least one of the following sequences compared to SEQ ID NO: 2666: F136W, S162C, S162E, S162F, S162H, S162T, S162V, K164A, K164S, K164W, K167A, K170T, R200C / H271G, K259V, and G373S.

[0175] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and has the following sequence identity compared to SEQ ID NO: 2666: 20, 20 / 100, 20 / 100 / 104 / 111 / 242, 20 / 100 / 104 / 120 / 197 / 367, 20 / 100 / 104 / 197 / 203 / 242 / 292, 20 / 100 / 104 / 203 / 350 / 353, 20 / 100 / 1 11, 20 / 100 / 120 / 197 / 232 / 235 / 315 / 366 / 367, 20 / 100 / 197 / 292 / 315, 20 / 100 / 203, 20 / 100 / 232 / 292 / 366 / 367, 20 / 100 / 235 / 315 / 367, 20 / 104 / 111 / 120 / 203, 20 / 120 / 197 / 292, 20 / 120 / 235, 20 / 292, 100, 100 / 111, 100 / 120 / 197 / 242, 104 / 120 / 232 / 353, and 111 / 197 / 242.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and has at least one of the following sequences compared to SEQ ID NO: 2666: 20E / 100D / 120I / 197E / 232M / 235M / 315V / 366S / 367K, 20K, 20K / 100D, 20K / 100D / 104L / 203V / 350L / 353D, 20K / 100D / 111R, 20K / 100D / 232M / 292K / 366S / 367K, 20K / 120I / 197E / 292K, 20K / 1 20I / 235M, 20K / 292K, 20R / 100D / 104L / 111R / 242R, 20R / 100D / 104L / 120I / 197E / 367K, 20R / 1 00D / 104L / 197E / 203V / 242R / 292K, 20R / 100D / 197E / 292K / 315V, 20R / 100D / 203V, 20R / 100D / 2 and amino acid sequences having one or more residue differences or sets of residue differences selected from 35M / 315V / 367K, 20R / 104L / 111R / 120I / 203V, 100D, 100D / 111R, 100D / 120I / 197E / 242R, 104L / 120I / 232M / 353D, and 111R / 197E / 242R.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2666, and the following sequence identity compared to SEQ ID NO: 2666: G20E / Q100D / V120I / N197 E / G232M / E235M / I315V / K366S / R367K, G20K, G20K / Q100D, G20K / Q100D / V104L / L203V / E350L / G353D, G20K / Q100D / E111R, G20K / Q100D / G232M / G292K / K366S / R367K, G20K / V120I / N197E / G292K, G20K / V120I / E235M, G20K / G292K, G20R / Q100D / V104L / E111R / K242R, G20R / Q100D / V104L / V120I / N197E / R367K, G20R / Q100D / V 104L / N197E / L203V / K242R / G292K, G20R / Q100D / N197E / G292K / I315V, G20R / Q100D / L203V, G20R / Q100D / E and amino acid sequences having one or more residue differences or sets of residue differences selected from 235M / I315V / R367K, G20R / V104L / E111R / V120I / L203V, Q100D, Q100D / E111R, Q100D / V120I / N197E / K242R, V104L / V120I / G232M / G353D, and E111R / N197E / K242R.

[0176] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:2794 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO:2794: 200, 201, 202, 203, 226, 229, 230, 234, 235, 236, 237, 238, 324, 326, 342, 344, 352, 355, 360, 366, 369, 371, 373, 374, 377, 378, 383, 388, and 390. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and has at least one of the following sequences compared to SEQ ID NO: 2794: 200A, 200M, 201R, 202V, 203A, 226L, 226M, 229S, 230T, 234H, 234L, 234M, 234S, 234T ... and 390Y.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and has at least one of the following sequences compared to SEQ ID NO: 2794: R200A, R200M, T201R, F202V, L203A, F226L, F226M, T229S, N230T, K234H, K234R, E235T, D23 6L, D237C, D237E, D237T, E238R, F324A, F324T, Q326S, R342L, R342V, L344M, R352L, R352V, I355L, A360G, K366E, K366Q, K366R, F369L, F369Y, K371A, G373A, G373S, G373T, S374N, E377D, I378V, G383N, E388A, E388L, and W390Y.

[0177] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 2794: 200, 203, 226, 234, 237, 342, 350, 360, 366, 369, 373, 377, 378, 380, and 390. In some embodiments, an engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 2794: 200A, 200G, 200M, 203A, 226L, 234R, 237C, 237E, 237R, 342L, 342V, 350I, 360R, 366E, 366Q, 369Y, 373A, 377D, 378V, 380M, and 390Y. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and has at least one of the following sequences compared to SEQ ID NO: 2794: R200A, R200B, R200C, R200D, R200E, R200F, R200G, R200H, R200I ... and W390Y, R342L, R342V, E350I, A360R, K366E, K366Q, F369Y, G373A, E377D, I378V, A380M, and W390Y.

[0178] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 2794: 200, 201, 203, 207, 226, 230, 233, 234, 235, 322, 342, 344, 352, 355, 360, 366, 371, 373, 374, 378, and 387. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and has at least one of the following sequences compared to SEQ ID NO: 2794: 200A, 200G, 200M, 201R, 203A, 204A, 204B, 204C, 204D, 204E, 204F, 204G, 204H, 204I, 204J, 204K ... and 387L.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and has at least one of the following sequences compared to SEQ ID NO: 2794: R200A, R200G, R200M, T201R, L203A, L207A, L207E, and an amino acid sequence having one or more residue differences or a set of residue differences selected from F226L, N230Y, R233S, K234H, K234R, E235T, T322C, T322P, R342L, R342V, L344M, L344T, R352L, R352V, I355L, A360G, K366E, K366Q, K371A, K371S, G373A, G373S, S374D, I378V, and V387L.

[0179] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and the following: 15, 200, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 28 and 390. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and the following sequences compared to SEQ ID NO: 2794: 15G, 200A, 200G, 200M, 200T, 200W, 202V, 203A, 205G, 206G, 207A, 207E, 207G, 226L, 226M, 229S, 230G ... , 230T, 231G, 233S, 234H, 234R, 234S, 237C, 237E, 237T, 321C, 324T, 326S, 327R, 330G, 342L, 344M, 344T, 349M, 349T, 352L, 353A, 353D, 353N, 360R, 366E, 369Y, 371A, 373A, 373T, 374D, 374N, 378V, 380M, 386F, 387L, 388L, 388Q, and 390Y.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and has at least one of the following sequences compared to SEQ ID NO: 2794: S15G, R200A, R200G, R200M, R200T, R200W, F202V, L203A, D205G, A206G, L207A, L207E, L207G, F226L, F226M, T229S, N230G, N230T, P23 1G, R233S, K234H, K234R, K234S, D237C, D237E, D237T, I321C, F324T, Q326S, Y327R, A330G, R342L, L344M, L344T, E349M, E349T, R352L, G353A, G353D, G353N, A360R, K366E, F369Y, K371A, G373A, G373T, S374D, S374N, I378V, A380M, Y386F, V387L, E388L, E388Q, and W390Y.

[0180] In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and the following sequences compared to SEQ ID NO: 2794: 55 / 80 / 174 / 268 / 355 / 366, 55 / 80 / 268 / 315 / 366, 55 / 80 / 268 / 346, 55 / 111 / 156 / 268 / 315 / 324 / 327 / 366 / 373, 55 / 111 / 268 / 346 / 355, 55 / 111 / 268 / 355 / 366, 55 / 111 / 315 / 355 / 373, 55 / 268, 55 / 268 / 315 / 346, 55 / 268 / 324 / 366, 55 / 268 / 346 / 355, 62 / 66 / 69 / 143 / 338 / 353, 62 / 66 / 100 / 101 / 104 / 203 / 235 / 338, 62 / 69 / 80 / 101 / 104 / 143 / 235 / 338, 62 / 203 / 211 / 235 / 338 / 350, 66 / 69 / 143 / 235 / 338, 69 / 80 / 203 / 211 / 278 / 338, 80 / 111 / 268 / 324 / 327 / 346 / 366 / 373, 80 / 111 / 355 / 366, 80 / 143 / 203 / 211 / 338, 80 / 268 / 315 / 346 / 355, 80 / 268 / 327 / 346 / 366, 80 / 268 / 346, 80 / 315 / 346 / 364 / 373, 80 / 346 / 366, 100 / 101 / 211 / 278 / 338 / 350 / 353, 11 and 327 / 346. In some embodiments, the engineered TdT polypeptide has an amino acid sequence that is at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 18397%, 98%, 99% or higher sequence identity, and the following compared to SEQ ID NO: 2794: 55L / 80S / 174Q / 268I / 355L / 366E, 55L / 80S / 268I / 315V / 366E, 55L / 80S / 268V / 346V, 55L / 111R / 156A / 268V / 315V / 324W / 327F / 366E / 373S, 55L / 111R / 268V / 346I / 355L, 55L / 111R / 268V / 355L / 366E, 55L / 111R / 315V / 355L / 373S, 55L / 268I / 346V / 355 L, 55L / 268V, 55L / 268V / 315V / 346V, 55L / 268V / 324W / 366E, 62E / 66Q / 69H / 143S / 338G / 353D, 62E / 66Q / 100D / 101C / 104L / 203V / 235M / 338G, 62E / 69H / 8 0S / 101C / 104L / 143S / 235M / 338G, 62E / 203V / 211A / 235M / 338G / 350L, 66Q / 69H / 143S / 235M / 338G, 69H / 80S / 203V / 211A / 278H / 338G, 80S / 111R / 268I / 3 24W / 327F / 346I / 366E / 373S, 80S / 111R / 355L / 366E, 80S / 143S / 203V / 211A / 338G, 80S / 268I / 315V / 346I / 355L, 80S / 268V / 327F / 346I / 366E, 80S / 268 V / 346I, 80S / 268V / 346V, 80S / 315V / 346A / 364P / 373S, 80S / 346A / 366E, 10 0D / 101C / 211A / 278H / 338G / 350L / 353D, 111R / 268I, 268I, 268I / 315V / 346V , 268I / 315V / 355L, 268I / 324W, 268I / 324W / 327F / 346V, 268V / 315V / 327F / 346V, 268V / 315V / 346A, 268V / 315V / 346I, 268V / 315V / 346V / 366E, 268V / 32 4W / 327F / 346I, 268V / 327F / 346A, 268V / 346A / 355L, 268V / 346I, 268V / 346 V, 268V / 355L / 366E, 315V / 324W / 327F / 355L / 366E, 324W / 346I / 355L / 366E,and 327F / 346A. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2794, and any of the following compared to SEQ ID NO: 2794: M55L / Y80S / L174Q / L268I / I355L / K366E, M55L / Y80S / L268I / I315V ... 8V / R346V, M55L / E111R / E156A / L268V / I315V / F324W / Y327F / K366E / G373S, M55L / E111R / L268V / R346I / I355L, M55L / E111R / L268V / I355L / K366E, M55L / E111R / I315V / I355L / G373S, M55L / L268I / R346V / I355L, M55L / L268V, M55L / L268V / I315V / R346V, M55L / L268V / F324W / K366E, K62E / I66Q / L69H / A143S / K338G / G353D, K62E / I66Q / Q100D / V101C / V104L / L203V / E235M / K338G, K62E / L69H / Y80S / V101C / V104L / A143S / E2 35M / K338G, K62E / L203V / T211A / E235M / K338G / E350L, I66Q / L69H / A143S / E235M / K338G, L69H / Y80S / L203V / T211A / N278H / K338G, Y80S / E11 1R / L268I / F324W / Y327F / R346I / K366E / G373S, Y80S / E111R / I355L / K366E, Y80S / A143S / L203V / T211A / K338G, Y80S / L268I / I315V / R346I / I355L, Y80S / L268V / Y327F / R346I / K366E, Y80S / L268V / R346I, Y80S / L268V / R346V, Y80S / I315V / R346A / L364P / G373S, Y80S / R346A / K366E,Q100D / V101C / T211A / N278H / K338G / E350L / G353D, E111R / L268I, L268I, L268I / I315V / R346V, L268I / I315V / I355L, L268I / F324W , L268I / F324W / Y327F / R346V, L268V / I315V / Y327F / R346V, L268V / I315V / R346A, L268V / I315V / R346I, L268V / I315V / R346V / K366 E, L268V / F324W / Y327F / R346I, L268V / Y327F / R346A, L268V / R346A / I355L, L268V / R346I, L268V / R346V, L268V / I355L / K366E, I315V / F324W / Y327F / I355L / K366E, F324W / R346I / I355L / K366E, and Y327F / R346A.

[0181] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2978, and the following sequences compared to SEQ ID NO: 2978: 62 / 69 / 100 / 235 / 268 / 346, 62 / 69 / 353, 62 / 111 / 235 / 315 / 355, 62 / 111 / 235 / 324 / 346, 62 / 235 / 268 / 327 / 346 / 350 / 353 / 355, 62 / 235 / 346 / 350 / 355, 62 / 315 / 327 / 353, 62 / 323 / 346 / 353 / 355, 66, 66 / 100 / 235 / 315 / 327 / 353 / 355, 66 / 111 / 346 / 353 / 355, 66 / 235 / 268 / 346, 66 / 235 / 346, 66 / 235 / 373, 69, 69 / 100, 69 / 100 / 111 / 298 / 353 / 355, 69 / 100 / 235, 69 / 100 / 353 / 366, 69 / 111 / 235, 69 / 111 / 235 / 300 / 353 / 355, 69 / 111 / 235 / 31 5, 69 / 235 / 315, 69 / 235 / 353, 69 / 268, 69 / 268 / 324 / 327 / 353 / 355, 69 / 268 / 346, 69 / 268 / 346 / 353, 69 / 315 / 353, 69 / 324 / 327 / 346, 69 / 324 / 346 / 350, 69 / 324 / 353, 69 / 353 / 355, 100, 100 / 111 / 353, 100 / 235 / 268 / 315 / 346, 100 / 235 / 268 / 346 / 355, 100 / 235 / 268 / 366 / 373, 100 / 235 / 346 / 350 / 353, 100 / 268 / 34 6, 100 / 268 / 366, 111 / 235 / 268 / 327 / 346, 111 / 235 / 268 / 346, 111 / 235 / 346 / 350, 111 / 268, 111 / 327, 232 / 346 / 350 / 355, 235 / 268 / 327 / 346, 235 / 268 / 346 , 235 / 315, 235 / 315 / 353 / 355, 235 / 346, 235 / 346 / 350, 235 / 353, 235 / 353 / 355, 268 / 346, 268 / 346 / 350 / 353 / 355, 268 / 353, 315 / 327 / 346, 324 / 327 / 346,In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having one or more residue differences or a set of residue differences selected from 324 / 346 / 350, 324 / 355, 327 / 346, 346, and 353 / 355. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2978, and any of the following compared to SEQ ID NO: 2978: 62E / 69H / 100D / 235M / 268V / 346I, 62E / 69H / 353D, 62E / 111R / 235M / 315V / 355L, 62E / 111R / 235M / 324W / 346I, 62E / 235M / 268V / 327F / 346I / 350L / 353D / 355L, 62E / 235M / 346V / 350L / 355L, 62E / 315V / 327F / 353D, 62E / 323S / 346I / 35 3D / 355L, 66Q, 66Q / 100D / 235M / 315V / 327F / 353D / 355L, 66Q / 111R / 346V / 353D / 355L, 66Q / 235M / 268V / 346I, 66Q / 235M / 346S, 66Q / 235M / 37 3S, 69H, 69H / 100D, 69H / 100D / 111R / 298I / 353D / 355L, 69H / 100D / 235M, 69H / 100D / 353D / 366E, 69H / 111R / 235M, 69H / 111R / 235M / 300K / 35 3D / 355L, 69H / 111R / 235M / 315V, 69H / 235M / 315V, 69H / 235M / 353D, 69H / 268V, 69H / 268V / 324W / 327F / 353D / 355L, 69H / 268V / 346I, 69H / 268 V / 346V / 353D, 69H / 315V / 353D, 69H / 324W / 327F / 346I, 69H / 324W / 346I / 350L, 69H / 324W / 353D, 69H / 353D / 355L, 100D, 100D / 111R / 353D, 1 00D / 235M / 268V / 315V / 346I, 100D / 235M / 268V / 346V / 355L, 100D / 235M / 268V / 366E / 373S, 100D / 235M / 346I / 350L / 353D, 100D / 268V / 346I,100D / 268V / 366E, 111R / 235M / 268V / 327F / 346V, 111R / 235M / 268V / 346I, 111R / 235M / 346V / 350L, 111R / 268V, 111R / 327F, 232 D / 346I / 350L / 355L, 235M / 268V / 327F / 346V, 235M / 268V / 346I, 235M / 315V, 235M / 315V / 353D / 355L, 235M / 346I, 235M / 346V, 23 and 353D / 355L. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 2978, and has at least one of the following sequences compared to SEQ ID NO: 2978: K62E / L69H / Q100D / E235M / L268V / R346I, K62E / L69H / G353D, K62E / E111R / E235M / I315V / I355L, K62E / E111R / E235M / F324W / R346I, K62E / E235M / L268V / Y327F / R3 46I / E350L / G353D / I355L, K62E / E235M / R346V / E350L / I355L, K62E / I315V / Y327F / G35 3D, K62E / P323S / R346I / G353D / I355L, I66Q, I66Q / Q100D / E235M / I315V / Y327F / G353D / I355L, I66Q / E111R / R346V / G353D / I355L, I66Q / E235M / L268V / R346I, I66Q / E235M / R3 46S, I66Q / E235M / G373S, L69H, L69H / Q100D, L69H / Q100D / E111R / V298I / G353D / I355L,L69H / Q100D / E235M, L69H / Q100D / G353D / K366E, L69H / E111R / E235M, L69H / E111R / E235M / T300K / G353D / I355L, L69H / E111R / E235M / I315V, L69H / E2 35M / I315V, L69H / E235M / G353D, L69H / L268V, L69H / L268V / F324W / Y327F / G353D / I355L, L69H / L268V / R346I, L69H / L268V / R346V / G353D, L69H / I315V / G353D, L69H / F324W / Y327F / R346I, L69H / F324W / R346I / E350L, L69H / F32 4W / G353D, L69H / G353D / I355L, Q100D, Q100D / E111R / G353D, Q100D / E235M / L268V / I315V / R346I, Q100D / E235M / L268V / R346V / I355L, Q100D / E235M / L 268V / K366E / G373S, Q100D / E235M / R346I / E350L / G353D, Q100D / L268V / R34 6I, Q100D / L268V / K366E, E111R / E235M / L268V / Y327F / R346V, E111R / E235 M / L268V / R346I, E111R / E235M / R346V / E350L, E111R / L268V, E111R / Y327F, G232D / R346I / E350L / I355L, E235M / L268V / Y327F / R346V, E235M / L268V / R 346I, E235M / I315V, E235M / I315V / G353D / I355L, E235M / R346I, E235M / R34 and G353D / I355L.

[0182] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074, and one or more residue differences or sets of residue differences selected from the following: 16, 17, 210, 234, 373, and 380 compared to SEQ ID NO: 3074. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 3074: 16M, 17R, 210L, 234A, 234Q, 373N, and 380S. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 3074: N16M, T17R, I210L, K234A, K234Q, G373N, and A380S.

[0183] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 3074: 16, 17, 18, 20, 21, 22, 24, 27, 28, 29, 32, 38, 44, 48, 77, 78, 78 / 127, 79, 80, 103, 106, 111, 115, 117, 119, 210, 232, 238, 272, and 373. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074, and has at least one of the following sequences compared to SEQ ID NO: 3074: 16M, 17G, 18L, 20A, 20G, 21N, 22C, 22P, 22Q, 24T, 27S, 28G, 29 and amino acid sequences having one or more residue differences or sets of residue differences selected from H, 29S, 32C, 32P, 38R, 44R, 44V, 48I, 77L, 77V, 78L, 78L / 127R, 79R, 80W, 103C, 103G, 103P, 103S, 106Q, 106R, 106Y, 111H, 111M, 111N, 111S, 111V, 115R, 117I, 119A, 210L, 232T, 238D, 272D, and 373N.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074, and the following amino acids compared to SEQ ID NO: 3074: N16M, T17G, P18L, K20A, K20G, E21N, V22C, V22P, V22Q, S24T, A27S, R28G, P29H, P29S, Q32C , Q32P, K38R, Y44R, Y44V, F48I, I77L, I77V, F78L, F78L / K127R, K79R, S80W, D103C, D103G, D103P, D103S, E106Q, E106R, E106Y, E111H, E111M, E111N, E111S, E111V, S115R, V117I, E119A, I210L, G232T, E238D, K272D, and G373N.

[0184] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074, and the following sequences compared to SEQ ID NO: 3074: 111 / 226, 111 / 226 / 321 / 344 / 346 / 369 / 387, 162 / 367 / and amino acid sequences having one or more residue differences or sets of residue differences selected from: 371, 201 / 202, 201 / 202 / 272 / 360, 202 / 233, 202 / 235 / 360 / 367 / 371, 207 / 235 / 327 / 360 / 371, 226, 226 / 268, 226 / 321 / 369, 226 / 366, 226 / 366 / 369 / 387, 233 / 367 / 371, 233 / 371, and 367 / 371. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074, and has at least one of the following sequences compared to SEQ ID NO: 3074: 111R / 226L, 111R / 226L / 321C / 344T / 346I / 369Y / 387L, 162T / 367G / 371S, 201G / 202 and 367G / 371S, 226L, 226L / 268V, 226L / 321C / 369Y, 226L / 366E, 226L / 366E / 369Y / 387L, 233S / 367G / 371S, 233S / 371S, and 367G / 371S.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074, and has at least one of the following sequences compared to SEQ ID NO: 3074: E111R / F226L, E111R / F226L / I321C / L344T / R346I / F369Y / V387L, S162T / R367G / K371S, T201G / F202V ... and R367G / K371S.

[0185] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and the following sequences compared to SEQ ID NO: 3302: 75, 75 / 233 / 344, 75 / 233 / 366, 104 / 200 / 207 / 237 / 344 / 373 / 387, 104 / 207 / 344 / 387, 106 / 182 / 203 / 226 / 235 / 342 / 346 , 182 / 201 / 203 / 226 / 234 / 235 / 346, 182 / 201 / 203 / 226 / 234 / 342 / 346, 182 / 201 / 203 / 226 / 235, 182 / 201 / 203 / 226 / 235 / 346, 182 / 201 / 203 / 226 / 342 / 346, 182 / 201 / 203 / 268 / 346, 182 / 201 / 226 / 234 / 342 / 346, 182 / 201 / 226 / 235 / 346, 182 / 201 / 226 / 268 / 346, 182 / 201 / 342, 182 / 203 / 226, 182 / 226 / 234 / 268 / 3 42 / 346, 182 / 226 / 234 / 346, 182 / 226 / 235, 182 / 226 / 235 / 268 / 346, 182 / 226 / 235 / 342, 182 / 226 / 235 / 346, 182 / 226 / 268 / 342 / 346, 182 / 226 / 342, 182 / 2 26 / 346, 182 / 342 / 346, 201 / 203 / 226 / 234 / 342 / 346, 201 / 203 / 226 / 234 / 346, 201 / 203 / 226 / 268 / 342, 201 / 203 / 226 / 346, 201 / 203 / 268 / 346, 201 / 226 / 23 4 / 342 / 346, 201 / 226 / 234 / 346, 201 / 226 / 235 / 342, 201 / 226 / 268 / 346, 203 / 226, 203 / 226 / 234 / 235, 203 / 226 / 234 / 235 / 346, 203 / 226 / 235 / 268 / 346, 203 / 226 / 235 / 342, 203 / 226 / 235 / 346, 203 / 226 / 342 / 346, 203 / 226 / 346, 207, 207 / 233 / 237 / 344 / 387, 207 / 387, 226, 226 / 235 / 268, 226 / 235 / 268 / 342 / 346,and amino acid sequences having one or more residue differences or sets of residue differences selected from 226 / 235 / 268 / 346, 226 / 235 / 342, 226 / 268 / 342, 226 / 268 / 342 / 346, 226 / 268 / 346, 226 / 342, 226 / 342 / 346, 226 / 346, 259 / 276 / 387, and 346. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and has at least one of the following sequences compared to SEQ ID NO: 3302: 75V, 75V / 233S / 344T, 75V / 233S / 366E, 104L / 200A / 207E / 237R / 34 4M / 373A / 387L, 104L / 207E / 344T / 387L, 106D / 182G / 203A / 226L / 235T / 342L / 346I, 182G / 201R / 203A / 226L / 234H / 235T / 346I, 1 82G / 201R / 203A / 226L / 234R / 235T / 346I, 182G / 201R / 203A / 226L / 234R / 342V / 346I, 182G / 201R / 203A / 226L / 235T, 182G / 201R / 203A / 226L / 235T / 346I, 182G / 201R / 203A / 226L / 342V / 346I, 182G / 201R / 203A / 268I / 346I, 182G / 201R / 226L / 234H / 342V / 346 I, 182G / 201R / 226L / 235T / 346I, 182G / 201R / 226L / 268V / 346I, 182G / 201R / 342V, 182G / 203A / 226L, 182G / 226L / 234R / 268I / 34 2V / 346I, 182G / 226L / 234R / 346I, 182G / 226L / 235T, 182G / 226L / 235T / 268V / 346I, 182G / 226L / 235T / 342L, 182G / 226L / 235T / 3 46I, 182G / 226L / 268I / 342V / 346I, 182G / 226L / 342V, 182G / 226L / 346I, 182G / 342L / 346I, 201R / 203A / 226L / 234H / 342V / 346I,201R / 203A / 226L / 234H / 346I, 201R / 203A / 226L / 268I / 342V, 201R / 203A / 226L / 346I, 201R / 2 03A / 268I / 346I, 201R / 226L / 234H / 346I, 201R / 226L / 234R / 342L / 346I, 201R / 226L / 235T / 342 L, 201R / 226L / 268I / 346I, 203A / 226L, 203A / 226L / 234H / 235T / 346I, 203A / 226L / 234R / 235T, 203A / 226L / 235T / 268V / 346I, 203A / 226L / 235T / 342L, 203A / 226L / 235T / 346I, 203A / 226L / 34 2V / 346I, 203A / 226L / 346I, 207E, 207E / 233S / 237R / 344M / 387I, 207E / 387L, 226L, 226L / 235T / 268I, 226L / 235T / 268V / 342L / 346I, 226L / 235T / 268V / 346I, 226L / 235T / 342V, 226L / 268I / 3 and amino acid sequences having one or more residue differences or sets of residue differences selected from 42L / 346I, 226L / 268I / 342V, 226L / 268I / 346I, 226L / 268V / 346I, 226L / 342V, 226L / 342V / 346I, 226L / 346I, 259R / 276L / 387L, and 346I. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and has one of the following sequences compared to SEQ ID NO: 3302: A75V, A75V / R233S / L344T, A75V / R233S / K366E ... V104L / R200A / L207E / D237R / L344M / G373A / V387L, V104L / L207E / L344T / V387L, E106D / M182G / V203A / F226L / E235T / R342L / R346I, M182G / T201R / V203A / F226L / K234H / E235T / R346I, M182G / T201R / V203A / F226L / K234R / E235T / R346I,M182G / T201R / V203A / F226L / K234R / R342V / R346I、M182G / T201R / V203A / F226L / E235T、M182G / T201R / V203A / F226L / E235T / R346I、M182G / T201R / V203A / F226L / R342V / R346I、M182G / T201R / V203A / L268I / R346I、M182G / T201R / F226L / K234H / R342V / R346I、M182G / T201R / F226L / E235T / R346I、M182G / T201R / F226L / L268V / R346I、M182G / T201R / R342V、M182G / V203A / F226L、M182G / F226L / K234R / L268I / R342V / R346I、M182G / F226L / K234R / R346I、M182G / F226L / E235T、M182G / F226L / E235T / L268V / R346I、M182G / F226L / E235T / R342L、M182G / F226L / E235T / R346I、M182G / F226L / L268I / R342V / R346I、M182G / F226L / R342V、M182G / F226L / R346I、M182G / R342L / R346I、T201R / V203A / F226L / K234H / R342V / R346I、T201R / V203A / F226L / K234H / R346I、T201R / V203A / F226L / L268I / R342V、T201R / V203A / F226L / R346I、T201R / V203A / L268I / R346I、T201R / F226L / K234H / R346I、T201R / F226L / K234R / R342L / R346I、T201R / F226L / E235T / R342L、T201R / F226L / L268I / R346I、V203A / F226L、V203A / F226L / K234H / E235T / R346I、V203A / F226L / K234R / E235T、V203A / F226L / E235T / L268V / R346I、V203A / F226L / E235T / R342L、V203A / F226L / E235T / R346I、V203A / F226L / R342V / R346I、V203A / F226L / R346I、L207E、L207E / R233S / D237R / L344M / V387I, L207E / V387L, F226L, F226L / E235T / L268I, F226L / E235T / L26 8V / R342L / R346I, F226L / E235T / L268V / R346I, F226L / E235T / R342V, F226L / L268I / R342L / R346I, F and an amino acid sequence having one or more residue differences or a set of residue differences selected from: 226L / L268I / R342V, F226L / L268I / R346I, F226L / L268V / R346I, F226L / R342V, F226L / R342V / R346I, F226L / R346I, K259R / M276L / V387L, and R346I.

[0186] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and has the following sequence identity compared to SEQ ID NO: 3302: 75 / 233 / 344, 75 / 233 / 366, 104 / 200 / 207 / 237 / 344 / 373 / 387, 104 / 207 / 344 / 387, 182 / 201 / 203 / 226 / 234 / 235 / 346, 182 / 201 / 203 / 226 / 234 / 342 / 346, 182 / 201 / 203 / 226 / 235, 182 / 201 / 203 / 226 / 235 / 346, 182 / 201 / 203 / 226 / 342 / 346, 182 / 201 / 203 / 268 / 346, 182 / 201 / 226 / 234 / 342 / 346, 182 / 201 / 226 / 235 / 346, 182 / 201 / 226 / 268 / 346, 182 / 201 / 342, 182 / 203 / 226, 182 / 226 / 234 / 268 / 342 / 346, 182 / 226 / 234 / 346, 182 / 226 / 235, 182 / 226 / 235 / 268 / 346, 182 / 226 / 235 / 342, 182 / 226 / 235 / 346, 182 / 226 / 268 / 342 / 346, 182 / 226 / 342, 182 / 226 / 346, 201 / 203 / 226 / 234 / 342 / 346, 201 / 203 / 226 / 346, 201 / 203 / 268 / 342, 201 / 203 / 268 / 346, 201 / 226 / 234 / 342 / 346, 201 / 226 / 234 / 346, 201 / 226 / 235 / 342, 201 / 226 / 268 / 346, 203 / 226, 203 / 226 / 234 / 235, 203 / 226 / 235 / 268 / 346, 203 / 226 / 235 / 346, 203 / 226 / 346, 207, 207 / 233 / 237 / 344 / 387, 207 / 387, 226, 226 / 234 / 342, 226 / 235 / 268, 226 / 235 / 268 / 346, 226 / 235 / 346, 226 / 268 / 342, 226 / 268 / 346, 226 / 342 / 346, 226 / 346,and 259 / 276 / 387. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and the following residue differences compared to SEQ ID NO: 3302: 75V / 233S / 344T, 75V / 233S / 366E, 104L / 200A / 207E / 237R / 344M / 373A / 387L, 104L / 207E / 344T / 387L , 182G / 201R / 203A / 226L / 234H / 235T / 346I, 182G / 201R / 203A / 226L / 23 4R / 235T / 346I, 182G / 201R / 203A / 226L / 234R / 342V / 346I, 182G / 201R / 2 03A / 226L / 235T, 182G / 201R / 203A / 226L / 235T / 346I, 182G / 201R / 203A / 226L / 342V / 346I, 182G / 201R / 203A / 268I / 346I, 182G / 201R / 226L / 234H / 342V / 346I, 182G / 201R / 226L / 235T / 346I, 182G / 201R / 226L / 268V / 34 6I, 182G / 201R / 342V, 182G / 203A / 226L, 182G / 226L / 234H / 346I, 182G / 2 26L / 234R / 268I / 342V / 346I, 182G / 226L / 234R / 346I, 182G / 226L / 235T, 182G / 226L / 235T / 268V / 346I, 182G / 226L / 235T / 342L, 182G / 226L / 235T / 346I, 182G / 226L / 268I / 342V / 346I, 182G / 226L / 342V, 182G / 226L / 346 I, 201R / 203A / 226L / 234H / 342V / 346I, 201R / 203A / 226L / 346I, 201R / 20 3A / 268I / 346I, 201R / 203A / 268V / 342V, 201R / 226L / 234H / 346I, 201R / 2 26L / 234R / 342L / 346I, 201R / 226L / 235T / 342L, 201R / 226L / 268I / 346I,203A / 226L, 203A / 226L / 234R / 235T, 203A / 226L / 235T / 268V / 346I, 203A / 226L / 235T / 346I, 203A / 22 6L / 346I, 207E, 207E / 233S / 237R / 344M / 387I, 207E / 387L, 226L, 226L / 234R / 342V, 226L / 235T / 268I, and 259R / 276L / 387L. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and has at least one of the following sequence identity compared to SEQ ID NO: 3302: A75V / R233S / L3 44T, A75V / R233S / K366E, V104L / R200A / L207E / D237R / L344M / G373A / V387L, V104L / L207E / L344T / V3 87L, M182G / T201R / V203A / F226L / K234H / E235T / R346I, M182G / T201R / V203A / F226L / K234R / E235T / R3 46I, M182G / T201R / V203A / F226L / K234R / R342V / R346I, M182G / T201R / V203A / F226L / E235T, M182G / T 201R / V203A / F226L / E235T / R346I, M182G / T201R / V203A / F226L / R342V / R346I, M182G / T201R / V203A / L 268I / R346I, M182G / T201R / F226L / K234H / R342V / R346I, M182G / T201R / F226L / E235T / R346I, M182G / T201R / F226L / L268V / R346I, M182G / T201R / R342V, M182G / V203A / F226L, M182G / F226L / K234H / R346I,M182G / F226L / K234R / L268I / R342V / R346I, M182G / F226L / K234R / R346I, M182G / F226L / E235T, M182G / F226L / E235T / L268V / R346I, M182G / F226L / E235T / R342L, M182G / F226L / E235T / R346I, M182G / F226L / L268I / R342V / R346I, M182G / F226L / R342V, M182G / F226L / R346I, T201R / V203A / F226L / K234H / R342V / R346I, T201R / V203A / F226L / R346I, T201R / V203A / L268I / R346I, T201R / V203A / L268V / R342V, T201R / F226L / K234H / R346I, T201R / F226L / K234R / R342L / R346I, T201R / F226L / E235T / R342L, T 201R / F226L / L268I / R346I, V203A / F226L, V203A / F226L / K234R / E235T, V203A / F226L / E235T / L268V / R346I, V203A / F226L / E 235T / R346I, V203A / F226L / R346I, L207E, L207E / R233S / D237R / L344M / V387I, L207E / V387L, F226L, F226L / K234R / R342V, F2 and K259R / M276L / V387L.

[0187] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and the following sequences compared to SEQ ID NO: 3302: 75 / 207 / 373 / 378, 104 / 200 / 207 / 237 / 344 / 373 / 387, 104 / 207 / 344 / 387, 182 / 201 / 203 / 226 / 234 / 235 / 346, 182 / 201 / 203 / 226 / 234 / 342 / 346, 182 / 201 / 203 / 226 / 235, 182 / 201 / 203 / 226 / 235 / 346, 182 / 201 / 203 / 226 / 342 / 346, 182 / 201 / 203 / 268 / 346, 182 / 201 / 226 / 234 / 342 / 346, 182 / 201 / 226 / 235 / 346, 182 / 201 / 226 / 268 / 346, 182 / 203 / 226, 182 / 226 / 234 / 268 / 342 / 346, 182 / 226 / 234 / 34 6, 182 / 226 / 235, 182 / 226 / 235 / 268 / 346, 182 / 226 / 235 / 342, 182 / 226 / 235 / 346, 182 / 226 / 268 / 342 / 346, 182 / 226 / 342, 182 / 226 / 346, 201 / 203 / 226 / 234, 201 / 203 / 226 / 346, 201 / 203 / 268 / 342, 201 / 203 / 268 / 346, 201 / 226 / 234 / 342 / 346, 201 / 226 / 268 / 346, 201 / 226 / 346, 203 / 226, 203 / 226 26 / 234 / 235, 203 / 226 / 234 / 235 / 346, 203 / 226 / 235 / 268 / 346, 203 / 226 / 235 / 346, 203 / 226 / 346, 207, 207 / 233 / 237 / 344 / 387, 207 / 387, 226, 226 / 234 / 235 / 346, 226 / 235 / 268, 226 / 235 / 268 / 346, 226 / 235 / 342, 226 / 235 / 346, 226 / 268 / 342, 226 / 268 / 346, 226 / 342, 226 / 342 / 346, 226 / 346,and 259 / 276 / 387. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and the following compared to SEQ ID NO: 3302: 75V / 207E / 373A / 378V, 104L / 200A / 207E / 237R / 344M / 373A / 387L, 104L / 207E / 344T / 387L, 182G / 207E ... 1R / 203A / 226L / 234H / 235T / 346I, 182G / 201R / 203A / 226L / 234R / 235T / 346I, 182G / 201R / 203A / 226L / 234R / 342V / 346I, 182G / 201R / 203A / 226L / 235T, 182G / 201R / 203A / 226L / 235T / 346I, 182G / 201R / 203A / 226L / 34 2V / 346I, 182G / 201R / 203A / 268I / 346I, 182G / 201R / 226L / 234H / 342V / 3 46I, 182G / 201R / 226L / 235T / 346I, 182G / 201R / 226L / 268V / 346I, 182G / 203A / 226L, 182G / 226L / 234H / 346I, 182G / 226L / 234R / 268I / 342V / 346 I, 182G / 226L / 234R / 346I, 182G / 226L / 235T, 182G / 226L / 235T / 268V / 3 46I, 182G / 226L / 235T / 342L, 182G / 226L / 235T / 346I, 182G / 226L / 268I / 342V / 346I, 182G / 226L / 342V, 182G / 226L / 346I, 201R / 203A / 226L / 234 H, 201R / 203A / 226L / 346I, 201R / 203A / 268I / 346I, 201R / 203A / 268V / 34 2V, 201R / 226L / 234R / 342L / 346I, 201R / 226L / 268I / 346I, 201R / 226L / 3 46I, 203A / 226L, 203A / 226L / 234H / 235T / 346I, 203A / 226L / 234R / 235T,203A / 226L / 235T / 268V / 346I, 203A / 226L / 235T / 346I, 203A / 226L / 346I, 207E, 207E / 233S / 237R / 34 4M / 387I, 207E / 387L, 226L, 226L / 234R / 235T / 346I, 226L / 235T / 268I, 226L / 235T / 268V / 346I, 226L / and 259R / 276L / 387L. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and has at least one of the following sequences compared to SEQ ID NO: 3302: A75V / L 207E / G373A / I378V, V104L / R200A / L207E / D237R / L344M / G373A / V387L, V104L / L207E / L344T / V38 7L, M182G / T201R / V203A / F226L / K234H / E235T / R346I, M182G / T201R / V203A / F226L / K234R / E235T / R346I, M182G / T201R / V203A / F226L / K234R / R342V / R346I, M182G / T201R / V203A / F226L / E235T, M182G / T201R / V203A / F226L / E235T / R346I, M182G / T201R / V203A / F226L / R342V / R346I, M182G / T2 01R / V203A / L268I / R346I, M182G / T201R / F226L / K234H / R342V / R346I, M182G / T201R / F226L / E235 T / R346I, M182G / T201R / F226L / L268V / R346I, M182G / V203A / F226L, M182G / F226L / K234H / R346I,M182G / F226L / K234R / L268I / R342V / R346I, M182G / F226L / K234R / R346I, M182G / F226L / E235T, M182G / F226L / E235T / L268V / R34 6I, M182G / F226L / E235T / R342L, M182G / F226L / E235T / R346I, M182G / F226L / L268I / R342V / R346I, M182G / F226L / R342V, M182G / F226L / R346I, T201R / V203A / F226L / K234H, T201R / V203A / F226L / R346I, T201R / V203A / L268I / R346I, T201R / V203A / L268V / R34 2V, T201R / F226L / K234R / R342L / R346I, T201R / F226L / L268I / R346I, T201R / F226L / R346I, V203A / F226L, V203A / F226L / K234H / E 235T / R346I, V203A / F226L / K234R / E235T, V203A / F226L / E235T / L268V / R346I, V203A / F226L / E235T / R346I, V203A / F226L / R346 I, L207E, L207E / R233S / D237R / L344M / V387I, L207E / V387L, F226L, F226L / K234R / E235T / R346I, F226L / E235T / L268I, F226L / E2 and K259R / M276L / V387L.

[0188] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and the following sequences compared to SEQ ID NO: 3302: 75 / 233 / 344, 75 / 233 / 366, 104 / 200 / 207 / 237 / 344 / 373 / 387, 104 / 207 / 344 / 387, 106 / 182 / 203 / 226 / 235 / 342 / 346, 182 / 201 / 203 / 226 / 234 / 235 / 346, 182 / 201 / 203 / 226 / 234 / 342 / 346, 182 / 201 / 203 / 226 / 235, 182 / 201 / 203 / 226 / 235 / 346, 182 / 201 / 203 / 226 / 342 / 346, 182 / 201 / 203 / 268 / 346, 182 / 201 / 226 / 234 / 342 / 346, 182 / 201 / 226 / 235 / 346, 182 / 201 / 226 / 268 / 346, 182 / 201 / 342, 182 / 203 / 226, 182 / 226 / 234 / 268 / 342 / 346, 182 / 226 / 234 / 346, 182 / 226 / 235, 182 / 226 / 235 / 268 / 346, 182 / 226 / 235 / 342, 182 / 226 / 235 / 346, 182 / 226 / 268 / 342 / 346, 182 / 226 / 342, 182 / 226 / 346, 182 / 342 / 346, 201 / 203 / 226 / 268 / 342, 201 / 203 / 226 / 346, 201 / 203 / 268 / 342, 201 / 203 / 268 / 346, 201 / 226 / 234 / 342 / 346, 201 / 226 / 235 / 342, 201 / 226 / 268 / 346, 201 / 226 / 346, 201 / 235 / 268 / 342 / 346, 203 / 226, 203 / 226 / 234 / 235, 203 / 226 / 234 / 235 / 346, 203 / 226 / 235 / 268 / 346, 203 / 226 / 235 / 346, 203 / 226 / 342 / 346, 203 / 226 / 346, 207, 207 / 233 / 237 / 344 / 387, 207 / 387, 226, 226 / 234 / 235 / 346, 226 / 234 / 342, 226 / 235 / 268, 226 / 235 / 268 / 346,and amino acid sequences having one or more residue differences or sets of residue differences selected from 226 / 235 / 342, 226 / 235 / 346, 226 / 268 / 342, 226 / 268 / 346, 226 / 342, 226 / 342 / 346, 226 / 346, and 259 / 276 / 387. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and has at least one of the following sequences compared to SEQ ID NO: 3302: 75V / 233S / 344T, 75V / 233S / 366E, 104L / 200A / 207E / 237R / 344M / 373A / 387L , 104L / 207E / 344T / 387L, 106D / 182G / 203A / 226L / 235T / 342L / 346I, 182G / 201R / 203A / 226L / 234H / 235T / 346I, 182G / 201R / 203A / 2 26L / 234R / 235T / 346I, 182G / 201R / 203A / 226L / 234R / 342V / 346I, 182G / 201R / 203A / 226L / 235T, 182G / 201R / 203A / 226L / 235T / 346I , 182G / 201R / 203A / 226L / 342V / 346I, 182G / 201R / 203A / 268I / 346I, 182G / 201R / 226L / 234H / 342V / 346I, 182G / 201R / 226L / 235T / 3 46I, 182G / 201R / 226L / 268V / 346I, 182G / 201R / 342V, 182G / 203A / 226L, 182G / 226L / 234H / 346I, 182G / 226L / 234R / 268I / 342V / 346I , 182G / 226L / 234R / 346I, 182G / 226L / 235T, 182G / 226L / 235T / 268V / 346I, 182G / 226L / 235T / 342L, 182G / 226L / 235T / 346I, 182G / 22 6L / 268I / 342V / 346I, 182G / 226L / 342V, 182G / 226L / 346I, 182G / 342L / 346I, 201R / 203A / 226L / 268I / 342V, 201R / 203A / 226L / 346I,201R / 203A / 268I / 346I, 201R / 203A / 268V / 342V, 201R / 226L / 234R / 342L / 346I, 201R / 226L / 235T / 342L, 201R / 226L / 268I / 346I, 201R / 226L / 346I, 201R / 235T / 268V / 342V / 346I, 203 A / 226L, 203A / 226L / 234H / 235T / 346I, 203A / 226L / 234R / 235T, 203A / 226L / 235T / 268V / 34 6I, 203A / 226L / 235T / 346I, 203A / 226L / 342V / 346I, 203A / 226L / 346I, 207E, 207E / 233S / 23 7R / 344M / 387I, 207E / 387L, 226L, 226L / 234H / 342L, 226L / 234R / 235T / 346I, 226L / 234R / 3 42V, 226L / 235T / 268I, 226L / 235T / 268V / 346I, 226L / 235T / 342V, 226L / 235T / 346I, 226L / 2 and 259R / 276L / 387L. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3302, and has at least one of the following sequences compared to SEQ ID NO: 3302: A75V / R233S / L344T, A75V / R233S / K366E, V1 04L / R200A / L207E / D237R / L344M / G373A / V387L, V104L / L207E / L344T / V387L, E106D / M182G / V203A / F226L / E235T / R 342L / R346I, M182G / T201R / V203A / F226L / K234H / E235T / R346I, M182G / T201R / V203A / F226L / K234R / E235T / R346I,M182G / T201R / V203A / F226L / K234R / R342V / R346I、M182G / T201R / V203A / F226L / E235T、M182G / T201R / V203A / F226L / E235T / R346I、M182G / T201R / V203A / F226L / R342V / R346I、M182G / T201R / V203A / L268I / R346I、M182G / T201R / F226L / K234H / R342V / R346I、M182G / T201R / F226L / E235T / R346I、M182G / T201R / F226L / L268V / R346I、M182G / T201R / R342V、M182G / V203A / F226L、M182G / F226L / K234H / R346I、M182G / F226L / K234R / L268I / R342V / R346I、M182G / F226L / K234R / R346I、M182G / F226L / E235T、M182G / F226L / E235T / L268V / R346I、M182G / F226L / E235T / R342L、M182G / F226L / E235T / R346I、M182G / F226L / L268I / R342V / R346I、M182G / F226L / R342V、M182G / F226L / R346I、M182G / R342L / R346I、T201R / V203A / F226L / L268I / R342V、T201R / V203A / F226L / R346I、T201R / V203A / L268I / R346I、T201R / V203A / L268V / R342V、T201R / F226L / K234R / R342L / R346I、T201R / F226L / E235T / R342L、T201R / F226L / L268I / R346I、T201R / F226L / R346I、T201R / E235T / L268V / R342V / R346I、V203A / F226L、V203A / F226L / K234H / E235T / R346I、V203A / F226L / K234R / E235T、V203A / F226L / E235T / L268V / R346I、V203A / F226L / E235T / R346I、V203A / F226L / R342V / R346I、V203A / F226L / R346I、L207E、L207E / R233S / D237R / L344M / V387I、L207E / V387L, F226L, F226L / K234H / R342L, F226L / K234R / E235T / R346I, F226L / K234R / R342V, F226L / E235T / L268I, F226L / E235T / L268V / R346I, F226L / E235T / R342V, F226L / E235T / R346I, F226L / L268 and K259R / M276L / V387L.

[0189] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3398, and the following sequences compared to SEQ ID NO: 3398: 29 / 77 / 104 / 234 / 271, 29 / 77 / 104 / 234 / 271 / 279 / 380, 29 / 77 / 182 / 207 / 271, 29 / 77 / 207 / 234 / 279 / 380, 29 / 77 / 2 34 / 271 / 279 / 368, 29 / 77 / 271, 29 / 104 / 234 / 271 / 279 / 380, 29 / 182 / 207 / 380, 29 / 182 / 234 / 271 / 380, 29 / 182 / 271 / 279 / 380, 29 / 234 / 279 / 380, 29 / 271 / 279, 29 / 271 / 279 / 368 / 380, 32 / 78 / 106 / 200 / 226 / 272 / 373, 32 / 78 / 106 / 226, 32 / 78 / 200 / 226, 32 / 78 / 200 / 226 / 235 / 272, 32 / 78 / 200 / 226 / 235 / 321 / 373, 32 / 78 / 226, 32 / 78 / 226 / 321, 32 / 78 / 226 / 373, 32 / 106 / 200 / 226, 32 / 106 / 200 / 226 / 235 / 373, 32 / 200 / 226, 32 / 200 / 226 / 321 / 373, 32 / 226, 77 / 182 / 279 / 380, 77 / 182 / 368, 77 / 234 / 271 / 279 / 380, 78 / 106, 78 / 106 / 226 / 321 / 373, 78 / 106 / 235 / 321 / 373, 78 / 200 / 226 / 321 / In some embodiments, the amino acid sequence comprises an amino acid sequence having one or more residue differences or sets of residue differences selected from: 373, 78 / 226 / 321, 104 / 182, 106 / 200 / 226 / 373, 106 / 226 / 235 / 373, 182 / 207 / 279 / 368, 182 / 234 / 380, 182 / 279 / 368 / 380, 200 / 226, 200 / 226 / 321 / 373, 200 / 226 / 373, 207 / 271 / 368 / 380, 207 / 380, 226, 226 / 235 / 272 / 373, 226 / 272 / 373, and 271 / 380.Engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher sequence identity to the reference sequence SEQ ID NO: 3398, and the following sequences compared to SEQ ID NO: 3398: 29H / 77V / 104L / 234P / 271P, 29H / 77V / 104L / 234R / 271P / 279H / 380G, 29H / 77V / 182G / 207E / 271Q, 29H / 77V / 207E / 234H / 279H / 380G , 29H / 77V / 234H / 271P / 279H / 368Q, 29H / 77V / 271P, 29H / 104L / 234R / 271Q / 2 79H / 380G, 29H / 182G / 207E / 380G, 29H / 182G / 234P / 271P / 380G, 29H / 182G / 27 1Q / 279H / 380G, 29H / 234H / 279H / 380G, 29H / 271Q / 279H, 29H / 271Q / 279H / 36 8Q / 380G, 32C / 78L / 106R / 200A / 226L / 373N, 32C / 78L / 200A / 226L, 32C / 78L / 2 00A / 226L / 235T / 272D, 32C / 78L / 226L / 321M, 32C / 78L / 226L / 373N, 32C / 106 Q / 200A / 226L / 235T / 373N, 32C / 106R / 200A / 226L, 32P / 78L / 106Q / 226L, 32P / 78L / 106R / 200A / 226L / 272D / 373N, 32P / 78L / 106R / 200A / 226L / 373A, 32P / 78L / 200A / 226L / 235T / 321M / 373N, 32P / 78L / 226L, 32P / 78L / 226L / 373N, 32P / 200A / 226L, 32P / 200A / 226L / 321M / 373N, 32P / 226L, 77V / 182G / 279H / 380G , 77V / 182G / 368Q, 77V / 234H / 271Q / 279H / 380G, 78L / 106Q, 78L / 106R / 226L / 3 21M / 373A, 78L / 106R / 226L / 321M / 373N, 78L / 106R / 235T / 321M / 373N, 78L / 2 00A / 226L / 321M / 373N, 78L / 226L / 321M, 104L / 182G, 106R / 200A / 226L / 373A,and 271P / 380G. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3398, and has at least one of the following sequence identity compared to SEQ ID NO: 3398: P29H / I77V / V104L / K234P / H271P, ... 9H / I77V / V104L / K234R / H271P / F279H / A380G, P29H / I77V / M182G / L207E / H271Q, P29H / I77V / L207E / K234H / F27 9H / A380G, P29H / I77V / K234H / H271P / F279H / I368Q, P29H / I77V / H271P, P29H / V104L / K234R / H271Q / F279H / A380 G, P29H / M182G / L207E / A380G, P29H / M182G / K234P / H271P / A380G, P29H / M182G / H271Q / F279H / A380G, P29H / K23 4H / F279H / A380G, P29H / H271Q / F279H, P29H / H271Q / F279H / I368Q / A380G, Q32C / F78L / E106R / R200A / F226L / G37 3N, Q32C / F78L / R200A / F226L, Q32C / F78L / R200A / F226L / E235T / K272D, Q32C / F78L / F226L / I321M, Q32C / F78L / F226L / G373N, Q32C / E106Q / R200A / F226L / E235T / G373N, Q32C / E106R / R200A / F226L, Q32P / F78L / E106Q / F226L,Q32P / F78L / E106R / R200A / F226L / K272D / G373N, Q32P / F78L / E106R / R200A / F226L / G373A, Q32P / F78L / R200A / F2 26L / E235T / I321M / G373N, Q32P / F78L / F226L, Q32P / F78L / F226L / G373N, Q32P / R200A / F226L, Q32P / R200A / F226L / I321M / G373N, Q32P / F226L, I77V / M182G / F279H / A380G, I77V / M182G / I368Q, I77V / K234H / H271Q / F279H / A380G, F78L / E106Q, F78L / E106R / F226L / I321M / G373A, F78L / E106R / F226L / I321M / G373N, F78L / E106R / E235T / I321M / G 373N, F78L / R200A / F226L / I321M / G373N, F78L / F226L / I321M, V104L / M182G, E106R / R200A / F226L / G373A, E106R / F226L / E235T / G373A, M182G / L207E / F279H / I368Q, M182G / K234R / A380G, M182G / F279H / I368Q / A380G, R200A / F22 6L, R200A / F226L / I321M / G373N, R200A / F226L / G373A, L207E / H271Q / I368Q / A380G, L207E / A380G, F226L, F226L / E235T / K272D / G373A, F226L / K272D / G373N, and H271P / A380G.

[0190] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 3074: 16, 201, 203, 230, 231, 234, 235, 238, 271, 273, 275, 279, 321, 344, 349, 368, 373, and 380. In some embodiments, an engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO: 3074: 16M, 201V, 203G, 203I, 230E, 231I, 234A, 234L, 234P, 234Q, 235C, 235V, 238V, 271P, 271Q, 273D, 275E, 279H, 321M, 344C, 349C, 368Q, 373N, 380G, and 380S. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3074, and has at least one of the following sequences compared to SEQ ID NO: 3074: N16M, T201V, V203G, V20 and an amino acid sequence having one or more residue differences or a set of residue differences selected from: 3I, N230E, P231I, K234A, K234L, K234P, K234Q, M235C, M235V, E238V, H271P, H271Q, I273D, A275E, F279H, I321M, L344C, E349C, I368Q, G373N, A380G, and A380S.

[0191] In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3488, and the following sequences compared to SEQ ID NO: 3488: 14 / 15 / 77 / 78, 14 / 15 / 77 / 78 / 200 / 201 / 226 / 230 / 290, 14 / 15 / 77 / 78 / 200 / 203 / 230 / 290, 14 / 15 / 77 / 78 / 203 / 226 / 230 / 290, 14 / 15 / 77 / 200 / 226 / 230, 14 / 15 / 78, 14 / 15 / 78 / 226 / 230, 14 / 15 / 200 / 230, 14 / 77 / 78 / 200 / 226 / 230 / 257, 14 / 77 / 78 / 226 / 230 / 257, 14 / 77 / 201 / 230, 14 / 78 / 200 / 201 / 226 / 290, 14 / 78 / 200 / 201 / 339, 14 / 78 / 200 / 226 / 230 / 339, 14 / 78 / 200 / 226 / 290 / 339, 14 / 78 / 200 / 230 / 339, 14 / 200 / 203 / 226 / 230, 14 / 2 00 / 203 / 226 / 230 / 257 / 339, 14 / 200 / 203 / 230 / 339, 14 / 201 / 203 / 226, 15 / 77, 15 / 77 / 78, 15 / 77 / 78 / 200 / 201 / 203 / 226 / 230, 15 / 77 / 200 / 201 / 203 / 226 / 23 0 / 290 / 339, 15 / 77 / 200 / 230, 15 / 77 / 200 / 230 / 290 / 339, 15 / 77 / 201 / 226 / 230 / 339, 15 / 77 / 226 / 230 / 290 / 339, 15 / 78 / 201, 15 / 78 / 201 / 203 / 230, 15 / 78 / 2 01 / 230 / 290, 15 / 78 / 203 / 226 / 230, 15 / 200, 15 / 201 / 203 / 226 / 230 / 257 / 339, 15 / 226 / 230 / 339, 15 / 230, 32, 32 / 103, 32 / 103 / 106 / 207 / 210 / 235 / 321 / 368 , 32 / 103 / 207 / 228 / 232 / 373, 32 / 103 / 210 / 273 / 321 / 373, 32 / 103 / 273, 32 / 106, 32 / 106 / 207 / 210 / 232, 32 / 106 / 210, 32 / 106 / 210 / 373, 32 / 106 / 235 / 321,32 / 106 / 235 / 368, 32 / 106 / 273 / 321 / 373, 32 / 207 / 210 / 273, 32 / 207 / 210 / 368 / 373, 32 / 207 / 235, 32 / 210 / 232 / 235 / 368, 32 / 210 / 273, 32 / 235 / 273, 77 / 78 / 200 / 226 / 339, 77 / 78 / 201 / 226 / 257 / 339, 77 / 78 / 203, 77 / 78 / 203 / 230 / 339, 77 / 200 / 201, 77 / 200 / 201 / 226 / 230 / 339, 77 / 200 / 203 / 230 / 257 / 339, 77 / 200 / 230, 78, 78 / 200 / 230, 78 / 203 / 230 / 290, 78 / 226 / 230, 78 / 226 / 290, 103 / 106 / 207 / 321, 103 / 106 / 210, 103 / 207 / 210 / 235 / 3 21, 103 / 210 / 232 / 235, 103 / 232 / 273, 106, 106 / 207 / 210 / 232 / 235 / 321 / 368 / 373, 106 / 207 / 235 / 321 / 368, 106 / 235 / 273 / 373, 106 / 273, 200, 200 / 203 / 226 / 230 / 339, 200 / 230 / 257, 201, 201 / 203, 201 / 230 / 257, 201 / 2 and amino acid sequences having one or more residue differences or sets of residue differences selected from 30 / 290, 203, 203 / 226 / 230, 203 / 226 / 230 / 257, 207 / 235, 207 / 235 / 368, 207 / 368 / 373, 207 / 373, 232 / 235, 232 / 235 / 273 / 368, 232 / 235 / 321, 235, 235 / 273, and 273. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3488, and the following sequences compared to SEQ ID NO: 3488: 14E / 15G / 77V / 78L, 14E / 15G / 77V / 78L / 200A / 201L / 226L / 230P / 290R, 14E / 15G / 77V / 78L / 200A / 203S / 230I / 290H, 14E / 15G / 77V / 78L / 203S / 226L / 230I / 290R,14E / 15G / 77V / 200A / 226L / 230P、14E / 15G / 78L、14E / 15G / 78L / 226L / 230P、14E / 15G / 200A / 230I、14E / 77V / 78L / 200A / 226L / 230P / 257T、14E / 77V / 78L / 226L / 230P / 257T、14E / 77V / 201L / 230I、14E / 78L / 200A / 201L / 226L / 290R、14E / 78L / 200A / 201L / 339I、14E / 78L / 200A / 226L / 230P / 339I、14E / 78L / 200A / 226L / 290R / 339I、14E / 78L / 200A / 230I / 339I、14E / 200A / 203S / 226L / 230I、14E / 200A / 203S / 226L / 230P / 257T / 339I、14E / 200A / 203S / 230P / 339I、14E / 201L / 203S / 226L、15G / 77V、15G / 77V / 78L、15G / 77V / 78L / 200A / 201L / 203S / 226L / 230P、15G / 77V / 200A / 201L / 203S / 226L / 230P / 290R / 339I、15G / 77V / 200A / 230P、15G / 77V / 200A / 230P / 290R / 339I、15G / 77V / 201L / 226L / 230I / 339I、15G / 77V / 226L / 230P / 290R / 339I、15G / 78L / 201L、15G / 78L / 201L / 203S / 230P、15G / 78L / 201L / 230P / 290R、15G / 78L / 203S / 226L / 230I、15G / 200A、15G / 201L / 203S / 226L / 230P / 257T / 339I、15G / 226L / 230I / 339I、15G / 230I、15G / 230P、32P、32P / 103P、32P / 103P / 106R / 207E / 210L / 235T / 321M / 368Q、32P / 103P / 207E / 228V / 232I / 373D、32P / 103P / 210L / 273P / 321M / 373D、32P / 103P / 273P、32P / 106R、32P / 106R / 207E / 210L / 232I、32P / 106R / 210L、32P / 106R / 210L / 373D、32P / 106R / 235T / 321M、32P / 106R / 235T / 368Q、32P / 106R / 273Q / 321M / 373D, 32P / 207E / 210L / 273Q, 32P / 207E / 210L / 368Q / 373D, 32P / 207E / 235T, 32P / 210L / 232I / 235T / 368Q, 32P / 2 10L / 273P, 32P / 235T / 273Q, 77V / 78L / 200A / 226L / 339I, 77V / 78L / 201L / 226L / 257T / 339I, 77V / 78L / 203S, 77V / 78L / 203S / 230P / 339I, 7 7V / 200A / 201L, 77V / 200A / 201L / 226L / 230P / 339I, 77V / 200A / 203S / 230P / 257T / 339I, 77V / 200A / 230P, 78L, 78L / 200A / 230P, 78L / 203 S / 230P / 290R, 78L / 226L / 230I, 78L / 226L / 230P, 78L / 226L / 290R, 103P / 106R / 207E / 321M, 103P / 106R / 210L, 103P / 207E / 210L / 235T / 32 1M, 103P / 210L / 232N / 235T, 103P / 232I / 273Q, 106R, 106R / 207E / 210L / 232I / 235T / 321M / 368Q / 373D, 106R / 207E / 235T / 321M / 368Q, 10 6R / 235T / 273P / 373D, 106R / 273Q, 200A, 200A / 203S / 226L / 230P / 339I, 200A / 230I / 257T, 201L, 201L / 203S, 201L / 230P / 257T, 201L / 230 and 232I / 235T / 321M, 232N / 235T, 232N / 235T / 273Q / 368Q, 232N / 235T / 321M, 235T, 235T / 273P, and 273P. In some embodiments, the engineered TdT polypeptide has a sequence identity that is at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 110%, 111%, 112%, 113%, 114%, 115%, 116%, 117%, 118%, 119%, 120%, 121%, 122%, 123%, 124%, 125%, 126%, 127%, 128%, 129%, 130%, 131%, 132%, 133%, 134%, 135%, 136%, 137%, 138%, 139%, 140%, 141%, 142%, 143%, 144%, 145%, 146%, 147%, 148%, 149%, 150%, 151%, 152%, 153%, 154%, 155%, 156%, 157%, 158%, 159%, 160%, 161%, 162%, 163%, 164%, 165%, 166%, 167%, 168%, 169%, 170%, 171%, 172%, 173%, 174%, 175%, 176%, 177%, 178%, 179%, 180%, 181, 182, 183%,98%, 99% or higher sequence identity, and the following compared to SEQ ID NO: 3488: P14E / S15G / I77V / F78L, P14E / S15G / I77V / F78L / R200A / T201L / F226L / N230P / Q290R, P14E / S15G / I77V / F78L / R200A / V203S / N230I / Q290H, P14E / S15G / I77V / F78L / V203S / F226L / N230I / Q290R, P14E / S15G / I77V / F78L, P 14E / S15G / F78L / F226L / N230P, P14E / S15G / R200A / N230I, P14E / I77V / F78 L / R200A / F226L / N230P / D257T, P14E / I77V / F78L / F226L / N230P / D257T, P14 E / I77V / T201L / N230I, P14E / F78L / R200A / T201L / F226L / Q290R, P14E / F78L / R200A / T201L / Q339I, P14E / F78L / R200A / F226L / N230P / Q339I, P14E / F78L / R200A / F226L / Q290R / Q339I, P14E / F78L / R200A / N230I / Q339I, P14E / R20 0A / V203S / F226L / N230I, P14E / R200A / V203S / F226L / N230P / D257T / Q339I, P14E / R200A / V203S / N230P / Q339I, P14E / T201L / V203S / F226L, S15G / I77V , S15G / I77V / F78L, S15G / I77V / F78L / R200A / T201L / V203S / F226L / N230P, S 15G / I77V / R200A / T201L / V203S / F226L / N230P / Q290R / Q339I, S15G / I77V / R200A / N230P, S15G / I77V / R200A / N230P / Q290R / Q339I, S15G / I77V / T201L / F226L / N230I / Q339I, S15G / I77V / F226L / N230P / Q290R / Q339I, S15G / F78L / T201L, S15G / F78L / T201L / V203S / N230P, S15G / F78L / T201L / N230P / Q290R,S15G / F78L / V203S / F226L / N230I、S15G / R200A、S15G / T201L / V203S / F226L / N230P / D257T / Q339I、S15G / F226L / N230I / Q339I、S15G / N230I、S15G / N230P、Q32P、Q32P / D103P、Q32P / D103P / E106R / L207E / I210L / E、 235T / I321M / I368Q、Q32P / D103P / L207E / I228V / G232I / G373D、Q32P / D103P / I210L / I273P / I321M / G373D、Q32P / D103P / I273P、Q32P / E106R、Q32P / E106R / L207E / I210L / G232I、Q32P / E106R / I210L、Q32P / E106R / I210L / G373D、Q32P / E106R / E235T / I321M、Q32P / E106R / E235T / I368Q、Q32P / E106R / I273Q / I321M / G373D、Q32P / L207E / I210L / I273Q、Q32P / L207E / I210L / I368Q / G373D、Q32P / L207E / E235T、Q32P / I210L / G232I / E235T / I368Q、Q32P / I210L / I273P、Q32P / E235T / I273Q、I77V / F78L / R200A / F226L / Q339I、I77V / F78L / T201L / F226L / D257T / Q339I、I77V / F78L / V203S、I77V / F78L / V203S / N230P / Q339I、I77V / R200A / T201L、I77V / R200A / T201L / F226L / N230P / Q339I、I77V / R200A / V203S / N230P / D257T / Q339I、I77V / R200A / N230P、F78L、F78L / R200A / N230P、F78L / V203S / N230P / Q290R、F78L / F226L / N230I、F78L / F226L / N230P、F78L / F226L / Q290R、D103P / E106R / L207E / I321M、D103P / E106R / I210L、D103P / L207E / I210L / E235T / I321M、D103P / I210L / G232N / E235T、D103P / G232I / I273Q、E106R、E106R / L207E / I210L / G232I / E235T / I321M / I368Q / G373D、E106R / L207E / E235T / I321M / I368Q、E106R / E235T / I273P / G373D、E106R / I273Q、R200A、R200A / V203S / F226L / N230P / Q339I、R200A / N230I / D257T、T201L、T201L / V203S, T201L / N230P / D257T, T201L / N230P / Q290R, V203S, V203S / F226L / N230I / D2 57T, V203S / F226L / N230P, L207E / E235T, L207E / E235T / I368Q, L207E / I368Q / G373D, L207 and an amino acid sequence having one or more residue differences or a set of residue differences selected from E235T, E235T / I273Q / I368Q, G232N / E235T / I321M, E235T, E235T / I273P, and I273P.

[0192] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3488, and the following sequences compared to SEQ ID NO: 3488: 14 / 15 / 77 / 78 / 200 / 201 / 226 / 230 / 290, 14 / 15 / 77 / 78 / 200 / 203 / 230 / 290, 14 / 15 / 77 / 78 / 203 / 226 / 230 / 290, 14 / 15 / 77 / 200 / 226 / 230, 14 / 15 / 78 / 226 / 230, 14 / 15 / 200 / 230, 14 / 77 / 78 / 200 / 226 / 230 / 257, 14 / 77 / 201 / 230, 14 / 78 / 200 / 226 / 230 / 339, 14 / 78 / 200 / 226 / 290 / 339, 14 / 200 / 203 / 226 / 230, 14 / 201 / 203 / 226, 15 / 77, 15 / 77 / 78 / 200 / 201 / 203 / 226 / 230, 15 / 77 / 200 / 230, 15 / 77 / 201 / 226 / 230 / 339, 1 5 / 78 / 201 / 203 / 230, 15 / 78 / 203 / 226 / 230, 15 / 230, 32, 32 / 103 / 106 / 207 / 210 / 235 / 321 / 368, 32 / 103 / 210 / 273 / 321 / 373, 32 / 103 / 273, 32 / 106 / 207 / 210 / 232, 32 / 106 / 210, 32 / 106 / 235 / 368, 32 / 207 / 210 / 368 / 373, 32 / 207 / 235, 32 / 210 / 232 / 273 / 321, 32 / 210 / 232 / 273 / 368 / 373, 32 / 210 / 273, 32 / 235 / 273, 77 / 78 / 200 / 226 / 339, 77 / 200 / 230, 78 / 203 / 230 / 290, 78 / 226 / 230, 78 / 226 / 290, 103 / 106 / 207 / 321, 103 / 106 / 210, 103 / 207 / 210 / 235 / 321, 103 / 210 / 232 / 235, 103 / 232 / 273, 106 / 207 / 235 / 321 / 368, 106 / 235 / 273 / 373, 201 / 203, 203 / 226 / 230, 207 / 235 / 368, 232 / 235, 232 / 235 / 321, 235,and 235 / 273. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3488, and one or more of the following compared to SEQ ID NO: 3488: 14E / 15G / 77V / 78L / 200A / 201L / 226L / 230P / 290R, 14E / 15G / 77V / 78L / 200A / 203S / 230I / 290H ... G / 77V / 78L / 203S / 226L / 230I / 290R, 14E / 15G / 77V / 200A / 226L / 230P, 14 E / 15G / 78L / 226L / 230P, 14E / 15G / 200A / 230I, 14E / 77V / 78L / 200A / 226L / 230P / 257T, 14E / 77V / 201L / 230I, 14E / 78L / 200A / 226L / 230P / 339I, 14 E / 78L / 200A / 226L / 290R / 339I, 14E / 200A / 203S / 226L / 230I, 14E / 201L / 2 03S / 226L, 15G / 77V, 15G / 77V / 78L / 200A / 201L / 203S / 226L / 230P, 15G / 7 7V / 200A / 230P, 15G / 77V / 201L / 226L / 230I / 339I, 15G / 78L / 201L / 203S / 230P, 15G / 78L / 203S / 226L / 230I, 15G / 230I, 32P, 32P / 103P / 106R / 207E / 210L / 235T / 321M / 368Q, 32P / 103P / 210L / 273P / 321M / 373D, 32P / 103P / 273P, 32P / 106R / 207E / 210L / 232I, 32P / 106R / 210L, 32P / 106R / 235T / 36 8Q, 32P / 207E / 210L / 368Q / 373D, 32P / 207E / 235T, 32P / 210L / 232I / 273P / 321M, 32P / 210L / 232N / 273Q / 368Q / 373D, 32P / 210L / 273P, 32P / 235T / 2 73Q, 77V / 78L / 200A / 226L / 339I, 77V / 200A / 230P, 78L / 203S / 230P / 290R,78L / 226L / 230I, 78L / 226L / 230P, 78L / 226L / 290R, 103P / 106R / 207E / 321M, 103P / 106R / 210L , 103P / 207E / 210L / 235T / 321M, 103P / 210L / 232N / 235T, 103P / 232I / 273Q, 106R / 207E / 235T / 3 and amino acid sequences having one or more residue differences or sets of residue differences selected from 21M / 368Q, 106R / 235T / 273P / 373D, 201L / 203S, 203S / 226L / 230P, 207E / 235T / 368Q, 232N / 235T, 232N / 235T / 321M, 235T, and 235T / 273P. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3488, and has at least one of the following sequence identity compared to SEQ ID NO: 3488: P14E / S15G / I77V / F78L / R200A / T201L / F226L / N230P / Q290R, P14E / S15G / I77V / F78L / R200A / V203S / N230I / Q290H, P14E / S15G / I7 7V / F78L / V203S / F226L / N230I / Q290R, P14E / S15G / I77V / R200A / F226L / N230P, P14E / S15G / F78L / F226L / N 230P, P14E / S15G / R200A / N230I, P14E / I77V / F78L / R200A / F226L / N230P / D257T, P14E / I77V / T201L / N230 I, P14E / F78L / R200A / F226L / N230P / Q339I, P14E / F78L / R200A / F226L / Q290R / Q339I, P14E / R200A / V203S / F226L / N230I, P14E / T201L / V203S / F226L, S15G / I77V, S15G / I77V / F78L / R200A / T201L / V203S / F226L / N23 0P, S15G / I77V / R200A / N230P, S15G / I77V / T201L / F226L / N230I / Q339I, S15G / F78L / T201L / V203S / N230P,S15G / F78L / V203S / F226L / N230I, S15G / N230I, Q32P, Q32P / D103P / E106R / L207E / I210L / E235T / I321M / I368Q, Q32P / D1 03P / I210L / I273P / I321M / G373D, Q32P / D103P / I273P, Q32P / E106R / L207E / I210L / G232I, Q32P / E106R / I210L, Q32P / E1 06R / E235T / I368Q, Q32P / L207E / I210L / I368Q / G373D, Q32P / L207E / E235T, Q32P / I210L / G232I / I273P / I321M, Q32P / I2 10L / G232N / I273Q / I368Q / G373D, Q32P / I210L / I273P, Q32P / E235T / I273Q, I77V / F78L / R200A / F226L / Q339I, I77V / R20 0A / N230P, F78L / V203S / N230P / Q290R, F78L / F226L / N230I, F78L / F226L / N230P, F78L / F226L / Q290R, D103P / E106R / L20 7E / I321M, D103P / E106R / I210L, D103P / L207E / I210L / E235T / I321M, D103P / I210L / G232N / E235T, D103P / G232I / I273Q , E106R / L207E / E235T / I321M / I368Q, E106R / E235T / I273P / G373D, T201L / V203S, V203S / F226L / N230P, L207E / E235T / I368Q, G232N / E235T, G232N / E235T / I321M, E235T, and E235T / I273P.

[0193] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3958 and one or more residue differences or sets of residue differences selected from the following as compared to SEQ ID NO: 3958: 53, 262, 264, 265, 266, 267, 270, 271, 272, 273, 275, 277, 321, 322, 324, 325, 339, 346, 347, 350, 355, 360, 370, 371, 376, 380, 383, 386, and 388. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3958, and has at least one of the following sequences compared to SEQ ID NO: 3958: 53T, 262R, 264E, 264S, 265P, 266D, 266T, 267G, 267S, 270N, 271G, 272R, 273P, 273V, 275G, 276T, 277T, 278T, 279T, 280T, 281T, 282R, 283T, 284T, 285T, 286T, 287T, 288T, 289T, 290T, 291T, 292R, 293T, 294T, 295T, 296T, 297T, 298T, 299T, 300T, 301T, 302T, 303T, 304T, 305T, 306T, 307T, 308T, 309T, 310T, 311T, 312T, 313T, 314T, 315T, 316T, 317T, 318T, 319T, 319T, 320T, 321T, 322R, 323T, and 388H.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3958, and has at least one of the following sequences compared to SEQ ID NO: 3958: E53T, T262R, V264E, V264S, E265P, G266D, G266T, Q267G, Q267S, S270N, P271G, K272R, I273P, I273V, A275G, A275K, A275Q, and E388H.

[0194] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3958, and has at least one of the following sequences compared to SEQ ID NO: 3958: 53, 219, 229, 262, 270, 272, 276, 278, 279, 280, 282, 284, 286, 288, 289, 290, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, and amino acid sequences having one or more residue differences or sets of residue differences selected from 264, 265, 266, 267, 270, 271, 272, 273, 275, 277, 278, 280, 321, 322, 324, 325, 339, 346, 347, 350, 355, 360, 370, 371, 376, 380, 383, 386, 387, 388, and 390. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher sequence identity to the reference sequence SEQ ID NO: 3958, and any of the following sequences compared to SEQ ID NO: 3958: 53T, 219T, 229V, 262R, 264E, 265P, 266D, 266T, 267E, 267G, 267S, 270N, 271G, 271T, 272R, 273P, 273V, 275G, 275K, 275Q, 275R, 275S, 277S, 278G, 279G, 280G, 281G, 282G, 283G, 284G, 285G, 286G, 287G, 288G, 289G, 290G, 291G, 292G, 293G, 294G, 295G, 296G, 297G, 298G, 299G, 300G, 301G, 302G, 303G, 304G, 305G, 306G, 307G, 308G, 309G, 310G, 311G, 312G, 313G, 314G, 315G, 316G, 317G, 318G, 31 , 278P, 280S, 321C, 321S, 321V, 322A, 324A, 324D, 324E, 324L, 324P, 324R, 324V, 325A, 325G, 325H, 325K, 325P, 325R, 325V, 339M, 346L, 346Q, 347F, 347W, 350W, 355P, 360N, 360S, 370Y, 371E, 371G, 376A, 376R, 376T, 380S, 380T, 383D, 383E, 383T, 383V, 386W, 387I, 388H, and 390A.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3958, and :E53T, P219T, T229V, T262R, V264E, E265P, G266D, G266T, Q267E, Q267G, Q267S, S270N, P271 G, P271T, K272R, I273P, I273V, A275G, A275K, A275Q, A275R, A275S, D277S, N278G, N278P, Q28 0S, I321C, I321S, I321V, T322A, F324A, F324D, F324E, F324L, F324P, F324R, F324V, E325A, E 325G, E325H, E325K, E325P, E325R, E325V, Q339M, R346L, R346Q, Y347F, Y347W, E350W, L355P, and an amino acid sequence having one or more residue differences or a set of residue differences selected from R360N, R360S, L370Y, S371E, S371G, E376A, E376R, E376T, G380S, G380T, G383D, G383E, G383T, G383V, Y386W, L387I, E388H, and W390A.

[0195] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3958 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO: 3958: 72, 264, 267, 270, 271, 272, 273, 280, 321, 322, 324, 325, 339, 346, 347, 349, 355, 360, 371, and 383. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3958, and has at least one of the following sequences compared to SEQ ID NO: 3958: 72R, 264E, 264Q, 264S, 267M, 270N, 271R, 271E, 271F, 271G, 271H, 271I, 271J, 271K ... and amino acid sequences having one or more residue differences or sets of residue differences selected from 71G, 272R, 273P, 273V, 280S, 321S, 321V, 322A, 324D, 324E, 324L, 324R, 324V, 325K, 325R, 339M, 346S, 347F, 349A, 349S, 349V, 355P, 360G, 360S, 360T, 371R, 383D, and 383E.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3958, and has at least one of the following sequences compared to SEQ ID NO: 3958: K72R, V264E, V264Q, V264S, Q267M, S270N, P271G, K272R, and an amino acid sequence having one or more residue differences or a set of residue differences selected from I273P, I273V, Q280S, I321S, I321V, T322A, F324D, F324E, F324L, F324R, F324V, E325K, E325R, Q339M, R346S, Y347F, E349A, E349S, E349V, L355P, R360G, R360S, R360T, S371R, G383D, and G383E.

[0196] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3788, and the following sequences compared to SEQ ID NO: 3788: 32 / 103 / 207 / 210 / 344 / 349, 32 / 103 / 207 / 321 / 344, 32 / 207 / 210 / 279, 32 / 273 / 344 / 373, 32 / 279, 32 / 279 / 321, 104 / 106, 104 / 106 / 111 / 200 / 201 / 235 / 268 / 368, 104 / 106 / 200 / 201 / 268, 106 / 111 / 200 / 235 / 368, 106 / 200 / 201, 106 / 200 / 201 / 209 / 368, 106 / 200 / 201 / 368, 106 / 201 / 235, 111, 200 / 235 / 368, 201 / 368, 207 / 210 / 273 / 279 / 344, 207 / 210 / 273 / 321 / 344 / 349, 207 / 273, 210, 210 / 273 / 279, 210 / 279, 273 / 279, and 273 / 279 / 373.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3788, and has at least the following sequence identity compared to SEQ ID NO: 3788: 3P / 207E / 210L / 344T / 349S, 32P / 103P / 207E / 321M / 344T, 32P / 207E / 210L / 279H, 32P / 273P / 344T / 373D, 32P / 279H, 32P / 279H / 321M, 104L / 106Q / 111V / 200A / 201L / 235T / 268I / 368Q, 104L / 106R, 1 04L / 106R / 200A / 201R / 268I, 106Q / 200A / 201R / 368Q, 106Q / 201L / 235T, 106R / 111V / 200A / 235T / 368Q, 106R / 200A / 201L, 106R / 200A / 201R / 209A / 368Q, 111V, 200A / 235T / 368Q, 201L / 368Q, 207E / and 273P / 279H / 373D.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3788, and the following sequences compared to SEQ ID NO: 3788: Q32P / D103P / L207E / I210L / L344T / E349S, Q32P / D103P / L207E / I321M / L344T, Q32P / L207E / I210L / F279H, Q32P / I273P / L344T / G373D, Q32P / F279 H, Q32P / F279H / I321M, V104L / E106Q / E111V / R200A / T201L / E235T / L268I / I368Q, V104L / E106R, V104L / E106R / R200A / T201R / L268I, E106Q / R200A / T201R / I368Q, E106Q / T201L / E235T, E106R / E111V / R200A / E235T / I368Q, E106R / R200A / T201L, E106R / R200A / T201R / T209A / I368Q, E111V, R200A / E235T / I368Q, T201L / I368Q, L207E / I and I273P / F279H / G373D.

[0197] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3788, and the following sequences compared to SEQ ID NO: 3788: 32 / 103 / 207 / 210 / 344 / 349, 32 / 103 / 207 / 321 / 34 ... 03 / 321 / 344, 32 / 207 / 210 / 279, 32 / 207 / 273 / 279 / 321, 32 / 207 / 279, 32 / 207 / 344, 32 / 273 / 279 / 344 / 349, 32 / 273 / 344 / 373, 32 / 279, 32 / 279 / 321, 104 / 106, 104 / 106 / 111 / 200 / 201 / 235 / 268 / 368, 104 / 106 / 111 / 200 / 201 / 268, 104 / 106 / 111 / 201 / 2 35 / 368, 106, 106 / 111 / 200 / 201, 106 / 111 / 200 / 201 / 268 / 368, 106 / 111 / 200 / 235 / 368, 106 / 111 / 201, 106 / 111 / 201 / 368, 106 / 200 / 201, 106 / 200 / 201 / 235, 106 / 201 / 235, 106 / 201 / 268, 111, 111 / 200 / 201 / 268, 111 / 200 / 368, 200 / 235 / 368, 200 / 3 68, 207 / 210 / 273 / 279 / 344, 207 / 210 / 273 / 279 / 344 / 349 / 373, 207 / 210 / 273 / 321 / 344 / 349, 207 / 273, 207 / 279 / 349, 210, 210 / 273 / 279, 210 / 273 / 279 / 349, 210 / 279, 273 / 279, and 273 / 279 / 373. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3788, and the following sequences compared to SEQ ID NO: 3788: 32P / 103P / 207E / 210L / 344T / 349S,32P / 103P / 207E / 321M / 344T, 32P / 103P / 321M / 344T, 32P / 207E / 210L / 279H, 32P / 207E / 273P / 279H / 321M, 32P / 207E / 279H, 32P / 207E / 344T, 32P / 273P / 279H / 344T / 349S, 32P / 273P / 344T / 373D, 32P / 279H, 32P / 279H / 321M, 104L / 106Q / 111V / 200A / 201L / 235T / 268I / 368Q, 104L / 106R, 104L / 106R / 111V / 200A / 201L / 268I, 104L / 106R / 111V / 201R / 235T / 368Q, 106Q / 200A / 201R / 235T, 106Q / 201L / 235T, 106R, 106R / 111V / 200A / 201L, 106R / 111V / 200A / 201L / 268I / 368Q, 106R / 111V / 200A / 201R / 268I / 368Q, 106R / 111V / 200A / 235T / 368Q, 106R / 111V / 201L, 106R / 111V / 201R / 368Q, 106R / 200A / 201L, 106R / 201R / 268I, 111V, 111V / 200A / 201R / 268I, 111V / 200A / 368Q, 200A / 235T / 368Q, 200A / 368Q, 207E / 210L / 273P / 279H / 344T, 207E / 210L / and amino acid sequences having one or more residue differences or sets of residue differences selected from 273P / 279H / 344T / 349S / 373D, 207E / 210L / 273P / 321M / 344T / 349S, 207E / 273P, 207E / 279H / 349S, 210L, 210L / 273P / 279H, 210L / 273P / 279H / 349S, 210L / 279H, 273P / 279H, and 273P / 279H / 373D. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 3788, and the following sequence identity compared to SEQ ID NO: 3788: Q32P / D103P / L207E / I210L / L344T / E349S,Q32P / D103P / L207E / I321M / L344T, Q32P / D103P / I321M / L344T, Q32P / L2 07E / I210L / F279H, Q32P / L207E / I273P / F279H / I321M, Q32P / L207E / F279 H, Q32P / L207E / L344T, Q32P / I273P / F279H / L344T / E349S, Q32P / I273P / L344T / G373D, Q32P / F279H, Q32P / F279H / I321M, V104L / E106Q / E111V / R2 00A / T201L / E235T / L268I / I368Q, V104L / E106R, V104L / E106R / E111V / R 200A / T201L / L268I, V104L / E106R / E111V / T201R / E235T / I368Q, E106Q / R 200A / T201R / E235T, E106Q / T201L / E235T, E106R, E106R / E111V / R200A / T201L, E106R / E111V / R200A / T201L / L268I / I368Q, E106R / E111V / R200A / T201R / L268I / I368Q, E106R / E111V / R200A / E235T / I368Q, E106R / E111V / T201L, E106R / E111V / T201R / I368Q, E106R / R200A / T201L, E106R / T201R / L268I, E111V, E111V / R200A / T201R / L268I, E111V / R200A / I368Q, R200 A / E235T / I368Q, R200A / I368Q, L207E / I210L / I273P / F279H / L344T, L207 E / I210L / I273P / F279H / L344T / E349S / G373D, L207E / I210L / I273P / I321M / L344T / E349S, L207E / I273P, L207E / F279H / E349S, I210L, I210L / I273P / F279H, I210L / I273P / F279H / E349S, I210L / F279H, I273P / F279H, and I273P / F279H / G373D.

[0198] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and the following sequences compared to SEQ ID NO: 4124: 32, 32 / 111, 32 / 112, 32 / 113, 32 / 114, 32 / 115, 32 / 116, 32 / 117, 32 / 118, 32 / 119 ... 1 / 235 / 271 / 272 / 339, 32 / 111 / 235 / 339, 32 / 111 / 235 / 386, 32 / 235, 32 / 235 / 271 / 339, 32 / 235 / 272, 32 / 235 / 272 / 339, 32 / 235 / 272 / 386, 32 / 235 / 339 / 386, 32 / 271 / 339, 32 / 272, 68 / 106 / 200, 68 / 344 / 383, 74 / 106 / 270 / 344, 103 / 324, 106 / 200 / 321 / 322 / 383, 106 / 270 / 344, 111 / 235, 111 / 235 / 271 / 339, 111 / 235 / 272 / 339 / 386, 111 / 235 / 339, 111 / 272 / 339 / 386, 200, 200 / 270, 200 / 270 / 322 / 383, 200 / 344, 235, 235 / 271 / 272 / and amino acid sequences having one or more residue differences or sets of residue differences selected from: 339 / 386, 235 / 271 / 339, 235 / 271 / 339 / 386, 235 / 272, 235 / 272 / 339, 235 / 272 / 386, 235 / 339, 235 / 339 / 386, 270 / 322 / 344 / 383, 324, 325, 344, 344 / 370, and 370. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4124, and The following: 32P, 32P / 111V / 235T, 32P / 111V / 235T / 271G / 272R / 339T, 32P / 111V / 235T / 339M, 32P / 111V / 235T / 339N, 32P / 111V / 235T / 386W, 32P / 235T, 32P / 235T / 271G / 339T, 32P / 235T / 272R,32P / 235T / 272R / 339M, 32P / 235T / 272R / 386W, 32P / 235T / 339M / 386W, 32P / 235T / 339N / 386 W, 32P / 271G / 339M, 32P / 272R, 68G / 106R / 200A, 68G / 344T / 383D, 74V / 106R / 270N / 344T, 103 P / 324E, 106R / 200A / 321V / 322A / 383D, 106R / 270N / 344T, 111V / 235T, 111V / 235T / 271G / 33 9N, 111V / 235T / 272R / 339N / 386W, 111V / 235T / 339T, 111V / 272R / 339N / 386W, 200A, 200A / 27 0N, 200A / 270N / 322A / 383D, 200A / 344T, 235T, 235T / 271G / 272R / 339M / 386W, 235T / 271G / 3 39M / 386W, 235T / 271G / 339N, 235T / 272R, 235T / 272R / 339M, 235T / 272R / 339T, 235T / 272R / 3 and amino acid sequences having one or more residue differences or sets of residue differences selected from 86W, 235T / 339M, 235T / 339M / 386W, 235T / 339N, 235T / 339N / 386W, 270N / 322A / 344T / 383D, 324R, 325H, 344T, 344T / 370Y, and 370Y. In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and the following sequences compared to SEQ ID NO: 4124: Q32P, Q32P / E111V / E235T, Q32P / E111V / E235T / P271G / K 272R / Q339T, Q32P / E111V / E235T / Q339M, Q32P / E111V / E235T / Q339N, Q32P / E111V / E235T / Y386W, Q32P / E235T, Q32P / E235 T / P271G / Q339T, Q32P / E235T / K272R, Q32P / E235T / K272R / Q339M, Q32P / E235T / K272R / Y386W, Q32P / E235T / Q339M / Y386W,Q32P / E235T / Q339N / Y386W, Q32P / P271G / Q339M, Q32P / K272R, C68G / E106R / R200A, C68G / L344T / G383D, A74V / E106R / S270N / L344T, D103P / F324E, E106R / R200A / I321V / T322A / G383D, E106R / S27 0N / L344T, E111V / E235T, E111V / E235T / P271G / Q339N, E111V / E235T / K272R / Q339N / Y386W, E111V / E235T / Q339T, E111V / K272R / Q339N / Y386W, R200A, R200A / S270N, R200A / S270N / T322A / G383D, R 200A / L344T, E235T, E235T / P271G / K272R / Q339M / Y386W, E235T / P271G / Q339M / Y386W, E235T / P27 1G / Q339N, E235T / K272R, E235T / K272R / Q339M, E235T / K272R / Q339T, E235T / K272R / Y386W, E235T / Q339M, E235T / Q339M / Y386W, E235T / Q339N, E235T / Q339N / Y386W, S270N / T322A / L344T / G383D, F324R, E325H, L344T, L344T / L370Y, and L370Y.

[0199] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and the following sequences compared to SEQ ID NO: 4124: 32 / 111 / 235, 32 / 111 / 23 5 / 339, 32 / 111 / 235 / 339 / 386, 32 / 111 / 235 / 386, 32 / 235, 32 / 235 / 272 / 339, 32 / 235 / 272 / 386, 32 / 235 / 339 / 386, 32 / 339, 68 / 106 / 200, 68 / 106 / 321 / 322, 68 / 200, 68 / 200 / 270 / 321, 68 / 270 / 321 / 322, 68 / 344, 68 / 3 44 / 383, 74 / 106 / 270 / 344, 103 / 324, 106 / 200 / 321 / 322 / 383, 106 / 270 / 344, 111 / 235, 111 / 235 / 272 / 339 / 386, 111 / 235 / 339, 111 / 272 / 339 / 386, 200, 200 / 270, 200 / 270 / 322 / 383, 200 / 344, 229, 229 / 325, 235, 235 / 271 / 272 / 339, 235 / 272, 235 / 272 / 339, 235 / 272 / 386, 235 / 339, 235 / 339 / 386, 262, 262 / 325 / 349, 270 / 322 / 344 / 383, 278, 324, 325, 344, 344 / 370, 349, and 370. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4124, and Compared to the following: 32P / 111V / 235T, 32P / 111V / 235T / 339M, 32P / 111V / 235T / 339N, 32P / 111V / 235T / 339T / 386W, 32P / 111V / 235T / 386W, 32P / 235T, 32P / 235T / 272R / 339M, 32P / 235T / 272R / 386W,32P / 235T / 339M / 386W, 32P / 235T / 339N / 386W, 32P / 235T / 339T / 386W, 32P / 339N, 68G / 106R / 20 0A, 68G / 106R / 321V / 322A, 68G / 200A, 68G / 200A / 270N / 321V, 68G / 270N / 321V / 322A, 68G / 344T, 68G / 344T / 383D, 74V / 106R / 270N / 344T, 103P / 324E, 106R / 200A / 321V / 322A / 383D, 106R / 270N / 344T, 111V / 235T, 111V / 235T / 272R / 339N / 386W, 111V / 235T / 339T, 111V / 272R / 339N / 386W, 200 A, 200A / 270N, 200A / 270N / 322A / 383D, 200A / 344T, 229V, 229V / 325H, 235T, 235T / 271G / 272R / 3 39N, 235T / 272R, 235T / 272R / 339M, 235T / 272R / 339T, 235T / 272R / 386W, 235T / 339M, 235T / 339M / 386W, 235T / 339N, 235T / 339N / 386W, 262R, 262R / 325H / 349N, 270N / 322A / 344T / 383D, 278R, 324R, 325H, 344T, 344T / 370Y, 349N, and 370Y. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and the following sequences compared to SEQ ID NO: 4124: Q32P / E111V / E235T, Q32P / E111V / E235 T / Q339M, Q32P / E111V / E235T / Q339N, Q32P / E111V / E235T / Q339T / Y386W, Q32P / E111V / E235T / Y386W, Q32P / E235T , Q32P / E235T / K272R / Q339M, Q32P / E235T / K272R / Y386W, Q32P / E235T / Q339M / Y386W, Q32P / E235T / Q339N / Y386W,Q32P / E235T / Q339T / Y386W, Q32P / Q339N, C68G / E106R / R200A, C68G / E106R / I321V / T322A, C68G / R200A, C 68G / R200A / S270N / I321V, C68G / S270N / I321V / T322A, C68G / L344T, C68G / L344T / G383D, A74V / E106R / S27 0N / L344T, D103P / F324E, E106R / R200A / I321V / T322A / G383D, E106R / S270N / L344T, E111V / E235T, E111V / E235T / K272R / Q339N / Y386W, E111V / E235T / Q339T, E111V / K272R / Q339N / Y386W, R200A, R200A / S270N, R20 0A / S270N / T322A / G383D, R200A / L344T, T229V, T229V / E325H, E235T, E235T / P271G / K272R / Q339N, E235T / K272R, E235T / K272R / Q339M, E235T / K272R / Q339T, E235T / K272R / Y386W, E235T / Q339M, E235T / Q339M / Y38 6W, E235T / Q339N, E235T / Q339N / Y386W, T262R, T262R / E325H / E349N, S270N / T322A / L344T / G383D, N278R, F324R, E325H, L344T, L344T / L370Y, E349N, and L370Y.

[0200] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and the following sequences compared to SEQ ID NO: 4124: 9 / 302, 26, 44 / 193, 47, 54, 58, 62, 63, 64, 70, 89, 99, 110, 144, 153, 154, 155, 159, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221 and 309.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and has at least one of the following sequences compared to SEQ ID NO: 4124: 9D / 302S, 26A, 26F, 26Q, 26T, 44C / 193D, 47L, 54R, 54V, 58D, 58S, 62A, 62G , 63S, 64Y, 70S, 89M, 89T, 99G, 99N, 99Q, 99V, 110M, 110N, 110S, 110V, 144I, 153C, 153M, 154L, 155Y, 159R, 159W, 161P, 162A, 162F, 162G, 162I, 162P, 163V, 164C, 164M, 170F, 174R, 181L, 184T, 189G, 189R, 190I, 192L, 193A, 193C, 193D, 193G, 193K, 193Y, 194L, 22 0S, 220Y, 237C, 237P, 237S, 237T, 237Y, 238S, 241P, 243A, 244C, 244I, 245G, 246I, 246M, 248R, 249N, 250R, 252S, 258C, 258V, 260N , 275S / 291N, 284S, 289D, 291N, 291R, 293A, 293G, 293I, 293S, 293T, 294A, 294K, 294V, 295L, 295P, 295S, 296R, 300M, 300P, 300S, 3 and 309Y.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and has at least one of the following sequences compared to SEQ ID NO: 4124: G9D / K302S, 126A, 126F, 126Q, 126T, Y44C / L193D, K47L, I54R, I54V, N58D, N58S, K62A, K62G, E63S, N64Y, E70S, K89 M, K89T, D99G, D99N, D99Q, D99V, E110M, E110N, E110S, E110V, E144I, T153C , T153M, V154L, E155Y, A159R, A159W, K161P, S162A, S162F, S162G, S162I, S 162P, L163V, K164C, K164M, K170F, L174R, S181L, S184T, D189G, D189R, A19 0I, T192L, L193A, L193C, L193D, L193G, L193K, L193Y, I194L, M220S, M220Y , D237C, D237P, D237S, D237T, D237Y, E238S, H241P, V243A, V244C, V244I, D 245G, L246I, L246M, K248R, K249N, Q250R, L252S, I258C, I258V, E260N, A27 5S / P291N, A284S, Y289D, P291N, P291R, V293A, V293G, V293I, V293S, V293T , D294A, D294K, D294V, N295L, N295P, N295S, S296R, T300M, T300P, T300S, T and V309Y.

[0201] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and the following sequences compared to SEQ ID NO: 4124: 9 / 302, 26, 55, 58, 62, 91, 110, 143, 148, 154, 162, 163, 170, 172, 174, 176, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 2 and 309.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and has at least one of the following sequences compared to SEQ ID NO: 4124: 9D / 302S, 26A, 26G, 26Q, 26T, 55V, 58D, 62R, 6 2W, 91L, 110L, 110M, 110N, 110T, 110V, 110Y, 143A, 148M, 154L, 162A, 162C, 162F, 162T, 162V, 163I, 173M, 174E, 184 L, 184Q, 184T, 190C, 190G, 192D, 193A, 193C, 193D, 193G, 193V, 193Y, 194C, 194L, 196A, 205A, 205E, 206P, 220S, 232N , 237C, 237L, 237P, 237S, 237T, 237Y, 238S, 245C, 245S, 246I, 248R, 249G, 249L, 249S, 253M, 275S / 291N, 284S, 286C , 287R, 289M, 289V, 291N, 291R, 291Y, 293I, 293S, 293T, 295L, 296C, 296R, 300F, 300M, 300P, 300V, 301A, 301G, 301T , 301V, 302A, 302C, 302L, 303A, 303G, 303M, 303N, 303P, 303R, 304G, 304L, 304V, 305G, 305S, 306A, 306G, 306L, 306P, 306T, 306W, 307S, 308L, 309A, 309D, 309F, 309L, 309N, and 309Y.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4124, and the following sequences compared to SEQ ID NO: 4124: G9D / K302S, I26A, I26G, I26Q, I26T, M55V, N58D, K62R, K62W, I91L, E110L, E110M , E110N, E110T, E110V, E110Y, S143A, R148M, V154L, S162A, S162C, S162F, S162T, S162V, L163I, F173M, L174E, S184L, S184Q, S184T, A190C, A190G, T192D, L193A, L193C, L193D, L193G, L193V, L193Y, I194C, I194L, K196A, D205A, D205E, A206P, M220S, G232N, D237C, D2 37L, D237P, D237S, D237T, D237Y, E238S, D245C, D245S, L246I, K248R, K249G, K249L, K249S, L253M, A275S / P291N, A284S, L286C, K28 7R, Y289M, Y289V, P291N, P291R, P291Y, V293I, V293S, V293T, N295L, S296C, S296R, T300F, T300M, T300P, T300V, S301A, S301G, S301T , S301V, K302A, K302C, K302L, K303A, K303G, K303M, K303N, K303P, K303R, S304G, S304L, S304V, D305G, D305S, M306A, M306G, M306L, M306P, M306T, M306W, A307S, E308L, V309A, V309D, V309F, V309L, V309N, and V309Y.

[0202] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4226, and and the following: 32 / 103 / 106 / 111 / 322 / 324, 32 / 106 / 280, 32 / 235, 32 / 235 / 386, 32 / 321 / 324 / 360, 32 / 324, 60 / 106 / 111 / 235 / 360, 60 / 280 / 360, 74 / 200 / 339, 103, 103 / 106 / 360, 103 / 111 / 235 / 280 / 360, 103 / 111 / 235 / 321 / 324, 106, 106 / 111, 106 / 111 / 322 / 383 / 386, 106 / 235, 106 / 235 / 383 / 386, 106 / 324, 111, 111 / 235 / 280, 111 / 322 / 360, 200 / 270 / 275 / 339, 235, 235 / 280, and amino acid sequences having one or more residue differences or sets of residue differences selected from 235 / 280 / 321 / 322 / 324, 235 / 280 / 321 / 324 / 383 / 386, 267 / 270 / 275 / 339 / 347, 267 / 272 / 275, 280, 280 / 383, 321 / 324, and 360 / 383.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and the following sequences compared to SEQ ID NO: 4226: 32P / 103P / 106R / 111V / 3 22T / 324E, 32P / 106R / 280S, 32P / 235T, 32P / 235T / 386W, 32P / 321V / 324D / 360S, 32P / 324D, 60R / 106R / 111V / 235T / 360S, 60R / 280S / 360S, 74V / 200A / 339M, 103P, 103P / 106R / 360S, 103P / 111V / 235T / 280S / 360S, 103 P / 111V / 235T / 321V / 324D, 106R, 106R / 111V, 106R / 111V / 322T / 383E / 386W, 106R / 235T, 106R / 235T / 383E / 386W, 106R / 324D, 111V, 111V / 235T / 280S, 111V / 322T / 360S, 200A / 270S / 275K / 339M, 235T, 235T / 280S, 2 and amino acid sequences having one or more residue differences or sets of residue differences selected from 35T / 280S / 321V / 322T / 324D, 235T / 280S / 321V / 324E / 383E / 386W, 267M / 270S / 275K / 339M / 347F, 267M / 272R / 275K, 280S, 280S / 383E, 321V / 324R, and 360S / 383E.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and the following sequences compared to SEQ ID NO: 4226: Q32P / D103P / E106R / E111V / A322T / F324E, Q32P / E106R / Q280S, Q32P / E235T, Q32P / E235T / Y386W, Q32P / I321V / F324D / R360S, Q32P / F324D, K60R / E106R / E111V / E235T / R360S, K 60R / Q280S / R360S, A74V / R200A / Q339M, D103P, D103P / E106R / R360S, D103P / E111V / E235T / Q280S / R360S, D103P / E111V / E 235T / I321V / F324D, E106R, E106R / E111V, E106R / E111V / A322T / D383E / Y386W, E106R / E235T, E106R / E235T / D383E / Y386 W, E106R / F324D, E111V, E111V / E235T / Q280S, E111V / A322T / R360S, R200A / N270S / A275K / Q339M, E235T, E235T / Q280S, E2 and amino acid sequences having one or more residue differences or sets of residue differences selected from 35T / Q280S / I321V / A322T / F324D, E235T / Q280S / I321V / F324E / D383E / Y386W, Q267M / N270S / A275K / Q339M / Y347F, Q267M / K272R / A275K, Q280S, Q280S / D383E, I321V / F324R, and R360S / D383E.

[0203] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:4226, and The following: 32 / 103 / 106 / 111 / 322 / 324, 32 / 235, 32 / 235 / 386, 32 / 321 / 324 / 360, 32 / 322 / 324 / 383 / 386, 32 / 324, 60 / 106 / 111 / 235 / 360, 72 / 74 / 200 / 272 / 339 / 347, 103, 103 / 106 / 360, 103 / 11 1 / 235 / 280 / 360, 103 / 111 / 235 / 321 / 324, 106, 106 / 111, 106 / 111 / 322 / 324 / 386, 106 / 111 / 322 / 383 / 386, 106 / 235, 106 / 235 / 383 / 386, 106 / 324, 111, 111 / 235 / 280, 111 / 322 / 360, 200 / 270 / 275 / 339, 235, 235 / 280, 235 / 280 / 321 / 322 / 324, 267 / 270 / 275 / 339 / 347, 267 / 272 / 275, 280, 280 / 383, 321 / 324, and 360 / 383.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and has at least one of the following sequences compared to SEQ ID NO: 4226: 32P / 103P / 106R / 111V / 32 2T / 324E, 32P / 235T, 32P / 235T / 386W, 32P / 321V / 324D / 360S, 32P / 322T / 324R / 383E / 386W, 32P / 324D, 60R / 106R / 111V / 235T / 360S, 72R / 74V / 200A / 272R / 339N / 347F, 103P, 103P / 106R / 360S, 103P / 111V / 235T / 280S / 360S, 103P / 111V / 235T / 321V / 324D, 106R, 106R / 111V, 106R / 111V / 322T / 324E / 386W, 106R / 111V / 322T / 38 3E / 386W, 106R / 235T, 106R / 235T / 383E / 386W, 106R / 324D, 111V, 111V / 235T / 280S, 111V / 322T / 360S, 200A / and amino acid sequences having one or more residue differences or sets of residue differences selected from 270S / 275K / 339M, 235T, 235T / 280S, 235T / 280S / 321V / 322T / 324D, 267M / 270S / 275K / 339M / 347F, 267M / 272R / 275K, 280S, 280S / 383E, 321V / 324R, and 360S / 383E.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and has one of the following sequences compared to SEQ ID NO: 4226: Q32P / D103P / E106R / E111V / A322T / F324E, Q32P / E235T, Q 32P / E235T / Y386W, Q32P / I321V / F324D / R360S, Q32P / A322T / F324R / D383E / Y386W, Q32P / F324D, K60R / E106R / E111V / E235 T / R360S, K72R / A74V / R200A / K272R / Q339N / Y347F, D103P, D103P / E106R / R360S, D103P / E111V / E235T / Q280S / R360S, D103 P / E111V / E235T / I321V / F324D, E106R, E106R / E111V, E106R / E111V / A322T / F324E / Y386W, E106R / E111V / A322T / D383E / Y3 86W, E106R / E235T, E106R / E235T / D383E / Y386W, E106R / F324D, E111V, E111V / E235T / Q280S, E111V / A322T / R360S, R200A / and amino acid sequences having one or more residue differences or sets of residue differences selected from N270S / A275K / Q339M, E235T, E235T / Q280S, E235T / Q280S / I321V / A322T / F324D, Q267M / N270S / A275K / Q339M / Y347F, Q267M / K272R / A275K, Q280S, Q280S / D383E, I321V / F324R, and R360S / D383E.

[0204] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and the following sequences compared to SEQ ID NO: 4226: 48, 50, 53, 57, 58, 63, 65, 68, 69, 73, 75, 85, 94, 104, 105, 123, 135, 140, 146, 150, 152, 154, 156, 158, 160, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 2 and amino acid sequences having one or more residue differences or sets of residue differences selected from 140, 141, 166, 167, 180, 184, 185, 187, 192, 209, 210, 217, 220, 239, 244, 253, 256, 260, 261, 265 / 346, 266, 267, 268, 273, 274, 275, 282, 284, 325, 344, 349, 350, 352, 353, 360, 361, 365, 371, 378, and 386.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and the following sequences compared to SEQ ID NO: 4226: 48I, 50E, 53D , 57L, 58D, 63D, 63G, 65G, 65N, 65R, 65V, 65W, 68M, 68R, 69A, 69H, 69M, 69T, 73K, 75G, 75I, 75V, 85I, 9 4V, 104I, 105L, 123N, 135I, 140I, 141R, 166N, 167R, 180G, 180R, 180V, 184D, 185F, 185R, 187D, 192L, 192Q, 209A, 210L, 210T, 210V, 217R, 220I, 220Q, 220R, 239V, 244L, 253I, 256G, 256Q, 260N, 261A, 26 1K, 261R, 265K / 346M, 266R, 267K, 268I, 273G, 274W, 275K, 275R, 275V, 282M, 284C, 284S, 325S, 344I, and an amino acid sequence having one or more residue differences or a set of residue differences selected from 344M, 349A, 349H, 349R, 349V, 349W, 349Y, 350I, 350R, 350V, 350Y, 352K, 353C, 360G, 360Q, 361C, 365Q, 371G, 371K, 371M, 371T, 371V, 378V, and 386F.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and has at least one of the following sequences compared to SEQ ID NO: 4226: F48I, D50E, E53D, E57L, N58D, E63D, E63G, L 65G, L65N, L65R, L65V, L65W, C68M, C68R, L69A, L69H, L69M, L69T, R73K, A75G, A75I, A75V, V85I, L94V, V104I, I105 L, D123N, V135I, V140I, K141R, S166N, K167R, A180G, A180R, A180V, S184D, K185F, K185R, E187D, T192L, T192Q, T20 9A, I210L, I210T, I210V, G217R, M220I, M220Q, M220R, L239V, V244L, L253I, E256G, E256Q, E260N, S261A, S261K, S 261R, E265K / R346M, G266R, Q267K, L268I, P273G, D274W, A275K, A275R, A275V, C282M, A284C, A284S, E325S, T344I, and an amino acid sequence having one or more residue differences or a set of residue differences selected from T344M, E349A, E349H, E349R, E349V, E349W, E349Y, E350I, E350R, E350V, E350Y, R352K, D353C, R360G, R360Q, L361C, K365Q, S371G, S371K, S371M, S371T, S371V, I378V, and Y386F.

[0205] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and the following: 48, 50, 53, 56, 57, 58, 63, 65, 68, 69, 70, 73, 75, 105, 135, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 410, 420, 430, 440, 450, 460, 470, 480, 490, 510, 520, 530, 540, 550, 560, 570, 580, 630, 650, 680, 690, 700, 730, 750, 1050, 1100, 1200, 1300, 1400, 1500, 1600, 1700, 1800, 1900, 2100, 2200, 2300, 2400, 2500, and amino acid sequences having one or more residue differences or sets of residue differences selected from 141, 166, 167, 175, 176, 180, 185, 187, 192, 210, 217, 220, 239, 244, 256, 260, 261, 266, 268, 271, 274, 275, 280, 282, 284, 325, 330, 344, 346, 349, 350, 352, 358, 360, 361, 369, 371, 378, 384, and 386.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and has at least one of the following sequences compared to SEQ ID NO: 4226: 48I, 48L, 50E, 53D, 53E, 53F, 53G, 53H, 53I, 53J, 53L ... 56V, 57L, 58D, 63D, 63G, 65A, 65G, 65N, 65R, 65V, 68M, 68Q, 68R, 68V, 69A, 69G, 69H, 69M, 69T, 70K, 70N, 70R, 73K, 75G, 75I, 75M, 75V, 105L, 135I, 140I, 141R, 166N, 167R, 175H, 176H, 180G, 180R, 180V, 185F, 1 85M, 185R, 187D, 192L, 192Q, 210L, 210V, 217R, 220I, 220Q, 220R, 239V, 244L, 256G, 256Q, 256S, 260N, 261A, 261H, 261K, 261R, 266R, 268I, 271R, 274G, 275K, 275R, 275V, 280G, 282M, 284C, 284S, 325S, 330T, and an amino acid sequence having one or more residue differences or a set of residue differences selected from 344I, 346K, 349A, 349R, 349V, 349W, 349Y, 350C, 350I, 350R, 350V, 350Y, 352K, 358A, 360G, 360K, 360Q, 361C, 369L, 371G, 371K, 371V, 378V, 384V, and 386F.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4226, and has at least one of the following sequences compared to SEQ ID NO: 4226: F48I, F48L, D50E, E53D, A56V, E57L, N58D, E63D, E 63G, L65A, L65G, L65N, L65R, L65V, C68M, C68Q, C68R, C68V, L69A, L69G, L69H, L69M, L69T, E70K, E70N, E70R, R73K, A75 G, A75I, A75M, A75V, I105L, V135I, V140I, K141R, S166N, K167R, Y175H, Y176H, A180G, A180R, A180V, K185F, K185M, K18 5R, E187D, T192L, T192Q, I210L, I210V, G217R, M220I, M220Q, M220R, L239V, V244L, E256G, E256Q, E256S, E260N, S261 A, S261H, S261K, S261R, G266R, L268I, P271R, D274G, A275K, A275R, A275V, Q280G, C282M, A284C, A284S, E325S, A330T, and Y386F.

[0206] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4734, and has at least one of the following sequences compared to SEQ ID NO: 4734: 32, 32 / 161 / 193 / 237 / 360, 32 / 162, 32 / 162 / 193 / 267, 32 / 162 / 237 / 265 / 266, 32 / 162 / 302, 32 / 302, 53 / 163 / 201 / 325 / 329, 53 / 200 / 201 / 325 / 329, 53 / 201, 53 / 201 / 275 / 2 80, 154, 154 / 166, 154 / 166 / 210 / 296, 154 / 339, 156 / 210 / 339, 161 / 162, 161 / 237, 162, 162 / 193 / 232 / 267 / 302 / 360, 162 / 237 / 265 / 266 / 267 / 302, 162 / 265, 162 / 267 / 360, 166 / 210 , 166 / 296, 166 / 346 / 347, 167 / 210 / 346 / 349, 193 / 267, 201 / 371, 210 / 339, 237, 237 / 265, 237 / 266 / 360, 237 / 360, 339, and 371.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4734, and the following sequences compared to SEQ ID NO: 4734: 32P, 32P / 161R / 193A / 23 7P / 360S, 32P / 162F, 32P / 162F / 193A / 267M, 32P / 162F / 237P / 265P / 266Q, 32P / 162F / 302L, 32P / 302L, 53T / 163M / 201R / 325H / 329F, 53T / 200A / 201R / 325H / 329F, 53T / 201R, 53T / 201R / 275K / 280S, 154R, 154R / 166N , 154R / 166T / 210L / 296R, 154R / 339M, 156V / 210L / 339M, 161G / 162F, 161G / 237P, 161R / 162F, 162F / 193A / 232N / 267M / 302L / 360S, 162F / 265P, 162F / 267M / 360S, 162R, 162R / 237P / 265P / 266Q / 267M / 302L, 166N / 21 0L, 166N / 296R, 166N / 346W / 347F, 167R / 210L / 346W / 349S, 193A / 267M, 201R / 371R, 210L / 339M, 237P, 237P / 265P, 237P / 266Q / 360S, 237P / 360S, 339M, and 371R.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4734, and has one of the following sequences compared to SEQ ID NO: 4734: Q32P, Q32P / K161R / L193A / D237P / R360S, Q32P / S162F ... Q32P / S162F / L193A / Q267M, Q32P / S162F / D237P / E265P / G266Q, Q32P / S162F / K302L, Q32P / K302L, E53T / L163M / T201R / E3 25H / Y329F, E53T / R200A / T201R / E325H / Y329F, E53T / T201R, E53T / T201R / A275K / Q280S, V154R, V154R / S166N, V154R / S1 66T / I210L / S296R, V154R / Q339M, E156V / I210L / Q339M, K161G / S162F, K161G / D237P, K161R / S162F, S162F / L193A / G232N / Q267M / K302L / R360S, S162F / E265P, S162F / Q267M / R360S, S162R, S162R / D237P / E265P / G266Q / Q267M / K302L, S166N / I2 10L, S166N / S296R, S166N / R346W / Y347F, K167R / I210L / R346W / E349S, L193A / Q267M, T201R / S371R, I210L / Q339M, D237P, D237P / E265P, D237P / G266Q / R360S, D237P / R360S, Q339M, and S371R.

[0207] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4734, and the following: 32, 32 / 162, 32 / 162 / 193, 32 / 162 / 193 / 267, 32 / 162 / 237 / 265 / 266, 32 / 162 / 302, 53 / 163 / 201 / 325 / 329, 53 / 200 / 201 / 325 / 329, 53 / 201, 53 / 201 / 275 / 280, 53 / 201 / 371, 82 / 154 / 296, 154, 154 / 166, 154 / 16 6 / 210 / 296, 154 / 166 / 346 / 347, 154 / 166 / 347, 154 / 167 / 210 / 347 / 349, 154 / 296 / 347, 154 / 339, 154 / 347, 156 / 166 / 167, 161 / 162, 161 / 237, 162, 162 / 193 / 232 / 267 / 302 / 360, 162 / 237 / 265 / 266 / 2 and amino acid sequences having one or more residue differences or sets of residue differences selected from 67 / 302, 162 / 265, 162 / 267 / 360, 163 / 201, 164 / 275 / 280, 166 / 210, 166 / 296, 166 / 346 / 347, 193 / 267, 201 / 371, 210 / 339, 237, 237 / 265, and 339.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4734, and has at least one of the following sequence identity compared to SEQ ID NO: 4734: 32P, 32P / 162F, 32P / 162F / 193A / 267M ... P / 162F / 237P / 265P / 266Q, 32P / 162F / 302L, 32P / 162R / 193A, 53T / 163M / 201R / 325H / 329F, 53T / 200A / 201R / 325H / 3 29F, 53T / 201R, 53T / 201R / 275K / 280S, 53T / 201R / 371R, 82S / 154R / 296R, 154R, 154R / 166N, 154R / 166N / 346S / 347F , 154R / 166N / 347F, 154R / 166T / 210L / 296R, 154R / 167R / 210L / 347F / 349S, 154R / 296R / 347F, 154R / 339M, 154R / 347 F, 156V / 166T / 167R, 161G / 237P, 161R / 162F, 162F / 193A / 232N / 267M / 302L / 360S, 162F / 265P, 162F / 267M / 360S, 16 and amino acid sequences having one or more residue differences or sets of residue differences selected from 2R, 162R / 237P / 265P / 266Q / 267M / 302L, 163M / 201R, 164M / 275K / 280S, 166N / 210L, 166N / 296R, 166N / 346W / 347F, 193A / 267M, 201R / 371R, 210L / 339M, 237P, 237P / 265P, and 339M.In some embodiments, the engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 4734, and the following sequences compared to SEQ ID NO: 4734: Q32P, Q32P / S162F, Q32P / S162F / L193A / Q267M, Q32P / S162F / D237P / E265P / G 266Q, Q32P / S162F / K302L, Q32P / S162R / L193A, E53T / L163M / T201R / E325H / Y329F, E53T / R200A / T201R / E325H / Y329F, E53T / T201R, E53T / T201R / A275K / Q280S, E53T / T201R / S371R, P82S / V154R / S296R, V154R, V154R / S166N, V154R / S166N / R346S / Y347F, V154R / S16 6N / Y347F, V154R / S166T / I210L / S296R, V154R / K167R / I210L / Y347F / E349S, V154R / S296R / Y347F, V154R / Q339M, V154R / Y347F, E15 6V / S166T / K167R, K161G / D237P, K161R / S162F, S162F / L193A / G232N / Q267M / K302L / R360S, S162F / E265P, S162F / Q267M / R360S, S16 2R, S162R / D237P / E265P / G266Q / Q267M / K302L, L163M / T201R, K164M / A275K / Q280S, S166N / I210L, S166N / S296R, S166N / R346W / Y347F, L193A / Q267M, T201R / S371R, I210L / Q339M, D237P, D237P / E265P, and Q339M.

[0208] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5052, and has at least one of the following sequences compared to SEQ ID NO: 5052: 32, 32 / 72 / 339, 32 / 72 / 339 / 360, 32 / 140 / 141 / 261 / 360, 32 / 140 / 339, 32 / 140 / 360, 32 / 141 / 180 / 244, 32 / 180 / 261 / 339, 32 / 244 / 261, 32 / 339, 53 / 73 / 75 / 237 / 23 9, 69 / 220, 69 / 339, 70, 70 / 72 / 180 / 360, 70 / 140 / 141 / 339, 70 / 360, 72 / 256 / 360, 72 / 360, 75 / 237 / 350, 140 / 141 / 256 / 261 / 339, 140 / 180 / 244 / 261 / 339 / 360, 141 / 244 / 261 / 36 and 261.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5052, and the following sequences compared to SEQ ID NO: 5052: 32P, 32P / 72R / 33 9M, 32P / 72R / 339M / 360G, 32P / 140I / 141R / 261R / 360S, 32P / 140I / 339M, 32P / 140I / 360G, 32P / 141R / 1 80R / 244L, 32P / 180R / 261R / 339M, 32P / 244L / 261R, 32P / 339M, 53T / 73K / 75V / 237P / 239V, 69A / 220R, 69 A / 339M, 70R, 70R / 72R / 180R / 360G, 70R / 140I / 141R / 339M, 70R / 360S, 72R / 256Q / 360S, 72R / 360G, 75I / 237P / 350R, 140I / 141R / 256Q / 261R / 339M, 140I / 180R / 244L / 261R / 339M / 360G, 141R / 244L / 261R / 360 and 261R, 261R, 261R / 339M, 261R, 261R / 360S, 261R / 339M, 261R, 261R / 360S, 261R, 261R / 339M, 261R ...In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5052, and has at least one of the following sequences compared to SEQ ID NO: 5052: Q32P, Q32P / K72R / Q339M, Q32P / K72R / Q339M / R 360G, Q32P / V140I / K141R / S261R / R360S, Q32P / V140I / Q339M, Q32P / V140I / R360G, Q32P / K141R / A180R / V244L, Q32P / A180R / S261R / Q339M, Q32P / V244L / S261R, Q32P / Q339M, E53T / R73K / A75V / D237P / L239V, L69A / M220R, L69A / Q339M, E 70R, E70R / K72R / A180R / R360G, E70R / V140I / K141R / Q339M, E70R / R360S, K72R / E256Q / R360S, K72R / R360G, A75I / D23 7P / E350R, V140I / K141R / E256Q / S261R / Q339M, V140I / A180R / V244L / S261R / Q339M / R360G, K141R / V244L / S261R / R36 and S261R / Q339M, K141R / V244L / R360S, K141R / E256Q / Q339M / R360G, K141R / S261R / Q339M, K141R / R360S, E155Y / M220R / E325S / Q339M, A180R / Q339M, M220R, M220R / Q339M, E256Q / R360S, and S261R.

[0209] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5052, and the following sequences compared to SEQ ID NO: 5052: 32 / 72 / 339, 32 / 72 / 339 / 360, 32 / 140 / 141 / 261 / 360, 32 / 140 / 339, 32 / 180 / 261 / 339, 32 / 244 / 261, 32 / 339, 53 / 73 / 75 / 237 / 239, 56 / 75 / 154 / 156 / 192 / 239 / 280 / 282, 56 / 75 / 192 / 239, 56 / 192 / 282 / 350, 65 / 220 / 339, 69 / 220, 69 / 339, 70 / 72, 70 / 72 / 140 / 244 / 261 / 339, 70 / 72 / 141 / 244, 70 / 261 / 339 / 360, 72 / 180 / 244 / 339, 72 / 256 / 360, 72 / 360, 140 / 141 / 256 / 261 / 339, 140 / 180 / 244 / 261 / 339 / 360, 141 / 244 / 360, 141 / 256 / 339 / 360, 1 and amino acid sequences having one or more residue differences or sets of residue differences selected from: 41 / 261 / 339, 141 / 360, 150 / 244 / 261 / 360, 154 / 156 / 282 / 350, 155 / 220 / 325 / 339, 180 / 339, 220, 220 / 339, 244 / 256 / 261 / 339 / 360, 325, 325 / 339, and 339.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5052, and has at least one of the following sequences compared to SEQ ID NO: 5052: 32P / 72R / 339M, 32P / 72R / 339M / 360G, 32P / 140I / 141R / 2 61R / 360S, 32P / 140I / 339M, 32P / 180R / 261R / 339M, 32P / 244L / 261R, 32P / 339M, 53T / 73K / 75V / 237P / 239V, 56V / 75I / 154V / 156V / 192L / 239V / 280G / 282M, 56V / 75V / 192L / 239V, 56V / 192Q / 282M / 350I, 65R / 220Q / 339M, 69A / 220R, 69A / 339M, 70R / 72 R, 70R / 72R / 140I / 244L / 261R / 339M, 70R / 72R / 141R / 244L, 70R / 261R / 339M / 360S, 72R / 180R / 244L / 339M, 72R / 256Q / 360S , 72R / 360G, 140I / 141R / 256Q / 261R / 339M, 140I / 180R / 244L / 261R / 339M / 360G, 141R / 244L / 360S, 141R / 256Q / 339M / 360G, and amino acid sequences having one or more residue differences or sets of residue differences selected from 141R / 261R / 339M, 141R / 360S, 150D / 244L / 261R / 360S, 154V / 156V / 282M / 350I, 155Y / 220R / 325S / 339M, 180R / 339M, 220R, 220R / 339M, 244L / 256Q / 261R / 339M / 360S, 325S, 325S / 339M, and 339M.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5052, and has at least one of the following sequence identity compared to SEQ ID NO: 5052: Q32P / K72R / Q339M, Q32P / K72R / Q339M / R360G, Q32P / V140I / K141R / S261R / R360S ... Q339M, Q32P / A180R / S261R / Q339M, Q32P / V244L / S261R, Q32P / Q339M, E53T / R73K / A75V / D237P / L239V, A56V / A75I / R154V / E156V / T192L / L23 9V / Q280G / C282M, A56V / A75V / T192L / L239V, A56V / T192Q / C282M / E350 I, L65R / M220Q / Q339M, L69A / M220R, L69A / Q339M, E70R / K72R, E70R / K7 2R / V140I / V244L / S261R / Q339M, E70R / K72R / K141R / V244L, E70R / S261R / Q339M / R360S, K72R / A180R / V244L / Q339M, K72R / E256Q / R360S, K72 R / R360G, V140I / K141R / E256Q / S261R / Q339M, V140I / A180R / V244L / S261R / Q339M / R360G, K141R / V244L / R360S, K141R / E256Q / Q339M / R360G, and amino acid sequences having one or more residue differences or sets of residue differences selected from K141R / S261R / Q339M, K141R / R360S, G150D / V244L / S261R / R360S, R154V / E156V / C282M / E350I, E155Y / M220R / E325S / Q339M, A180R / Q339M, M220R, M220R / Q339M, V244L / E256Q / S261R / Q339M / R360S, E325S, E325S / Q339M, and Q339M.

[0210] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:5152 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO:5152: 53, 53 / 237, 65 / 140, 65 / 140 / 192 / 193, 65 / 140 / 192 / 193 / 302, 65 / 193, 140 / 192 / 193 / 302, 140 / 193 / 302, 180, 192 / 193, 239, and 302. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5152, and the following sequences compared to SEQ ID NO: 5152: 53T, 53T / 237P ... and amino acid sequences having one or more residue differences or sets of residue differences selected from 65R / 140I, 65R / 140I / 192L / 193D, 65R / 140I / 192L / 193D / 302L, 65R / 193D, 140I / 192L / 193D / 302L, 140I / 193D / 302L, 180V, 192L / 193D, 239V, and 302L. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5152, and has one of the following sequences compared to SEQ ID NO: 5152: E53T, E53T / D237P, L65R / V140I, and amino acid sequences having one or more residue differences or sets of residue differences selected from L65R / V140I / T192L / L193D, L65R / V140I / T192L / L193D / K302L, L65R / L193D, V140I / T192L / L193D / K302L, V140I / L193D / K302L, A180V, T192L / L193D, L239V, and K302L.

[0211] In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence having at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:5252 and one or more residue differences or sets of residue differences selected from the following relative to SEQ ID NO:5252: 48 / 53 / 237 / 239, 48 / 256 / 261, 140, 141 / 261, 162, 220, 220 / 349, 261, 302, and 350. In some embodiments, the engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:5252 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO:5252: 48I / 53T / 237P / 239V, 48I / 256G / 261R, 140I, 141R / 261R, 162F, 220Q, 220Q / 349V, 261R, 302L, and 350Y. In some embodiments, an engineered TdT polypeptide comprises an amino acid sequence that has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:5252 and one or more residue differences or a set of residue differences selected from the following relative to SEQ ID NO:5252: F48I / E53T / D237P / L239V, F48I / E256G / S261R, V140I, K141R / S261R, S162F, M220Q, M220Q / E349V, S261R, K302L, and E350Y.

[0212] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5296, and the following sequences compared to SEQ ID NO: 5296: 65, 65 / 184 / 187, 82 / 184 / 220 / 386 , 154, 154 / 184, 154 / 187 / 220 / 350 / 386, 154 / 293, 154 / 350, 154 / 386, 184 / 187, 184 / 293, 187 / 220 / 350, 187 / 293, 220, 220 / 293, 293, 293 / 350, 293 / 350 / 386, 350, and 386. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5296, and has at least one of the following sequences compared to SEQ ID NO: 5296: 65R, 65R / 184T / 187D, 82S / 184T / 220Q / 386F, 154L, 154L / 187D, 154L / 184T / 187D, 154L / 184T / 187F, 154L / 184T / 187G ... and 386F.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5296, and has at least one of the following sequences compared to SEQ ID NO: 5296: L65R, L65R / S184T / E187D, P82S / S184T / M220Q / Y386F, R154L, R154L / S184T, R154L / E and Y386F, E350R, and Y386F.

[0213] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5296, and the following sequences compared to SEQ ID NO: 5296: 157, 160, 186, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, and amino acid sequences having one or more residue differences or sets of residue differences selected from 200, 201, 202, 203, 204, 206, 209, 210, 211, 227, 229, 230, 233, 234, 235, 236, 237 / 381, 238, 241, 242, 243, 247, 253, 258, 260, 267, 272, 278, 282, 286, 288, 290, 292, 297, 298, 299, 327, 331, 368, 373, and 381.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5296, and has at least one of the following sequences compared to SEQ ID NO: 5296: 157V, 160F, 160M, 160V, 160Y, 186A, 186L, 186V, 188K, 188M, 188V, 188Y, 186F, 186G, 186H, 186I, 186J, 186K, 186M, 186V, 186K, 186M, 186V, 186Y, 186K, 186M ... 9A, 189L, 189R, 190I, 190M, 190S, 191L, 191M, 192T, 192V, 192Y, 193E, 193G, 193Q, 193R, 194F, 194R, 194T, 194W, 195I, 195L, 19 6G, 196T, 197Q, 198V, 199M, 199W, 200L, 200M, 200T, 200W, 201S, 201Y, 202W, 203D, 203I, 203L, 203R, 204G, 206H, 209R, 210A, 210 C, 210T, 211V, 227V, 229G, 229R, 230N, 233L, 233S, 233W, 234G, 234Q, 234R, 234V, 235H, 235L, 236P, 237G / 381W, 238G, 238L, 238 R, 238W, 241A, 241E, 241L, 241R, 241S, 241W, 242V, 243M, 243S, 247V, 253V, 258C, 258L, 258V, 260A, 260G, 260M, 267T, 272R, 272S , 278S, 282T, 286A, 286C, 286V, 288A, 288K, 288Y, 290P, 292R, 297F, 297G, 297P, 297V, 298F, 298R, 298T, 299M, 299N, 299R, 299S, 299V, 299Y, 327H, 331K, 368G, 368Q, 368S, 368T, 368V, 373Q, 381L, and 381V.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5296, and has one of the following sequences compared to SEQ ID NO: 5296: A157V, S160F, S160M, S160V, S160Y, E186A, E186L, E186V, A188K, A188M, A188V, D189A, D189L, D189R, A190I, A190J, A190K, A190KM, A190K ... A190M, A190S, V191L, V191M, L192T, L192V, L192Y, D193E, D193G, D193Q, D193R, I194F, I194R, I194T, I194W, V195I, V195L, K196G, K196T, E197Q, T 198V, V199M, V199W, R200L, R200M, R200T, R200W, T201S, T201Y, V202W, S2 03D, S203I, S203L, S203R, P204G, A206H, T209R, I210A, I210C, I210T, A21 1V, L227V, T229G, T229R, I230N, R233L, R233S, R233W, P234G, P234Q, P23 4R, P234V, T235H, T235L, D236P, D237G / H381W, E238G, E238L, E238R, E238 W, H241A, H241E, H241L, H241R, H241S, H241W, K242V, V243M, V243S, L247V , L253V, I258C, I258L, I258V, E260A, E260G, E260M, Q267T, K272R, K272S, and an amino acid sequence having one or more residue differences or a set of residue differences selected from N278S, C282T, L286A, L286C, L286V, L288A, L288K, L288Y, R290P, K292R, A297F, A297G, A297P, A297V, V298F, V298R, V298T, G299M, G299N, G299R, G299S, G299V, G299Y, F327H, L331K, I368G, I368Q, I368S, I368T, I368V, D373Q, H381L, and H381V.

[0214] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5296, and the following sequences compared to SEQ ID NO: 5296: 160, 186, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 32, 33, 36, 37, 38, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 13 In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5296, and :160F, 160V, 160Y, 186A, 186L, 186V, 188K, 188M, 189A, 189R, 190I, 190M, 190Q, 190S, 191L, 191M, 192V, 192Y, 193E, 193G, 193Q, 194F, 195I, 196E, 196T, 197S, 199M, 199W, 200L, 200T, 2 00W, 201M, 202L, 203G, 204C, 204L, 209R, 209S, 210A, 210C, 210T, 211V, 229R, 229V, 230N, 23 3L, 234Q, 234V, 235H, 236R, 237G / 381W, 238L, 238R, 238W, 241R, 241S, 243M, 243S, 244S, 247 and an amino acid sequence having one or more residue differences or a set of residue differences selected from F, 253V, 260A, 260G, 278S, 286C, 286V, 288A, 292R, 297V, 298F, 298T, 299N, 299S, 327H, 331K, 368E, 368S, 368T, 373Q, 373R, and 381L.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5296, and has at least one of the following sequences compared to SEQ ID NO: 5296: S160F, S160V, S160Y, E 186A, E186L, E186V, A188K, A188M, D189A, D189R, A190I, A190M, A190Q, A190S, V191L, V191M, L192V, L19 2Y, D193E, D193G, D193Q, I194F, V195I, K196E, K196T, E197S, V199M, V199W, R200L, R200T, R200W, T201M, V202L, S203G, P204C, P204L, T209R, T209S, I210A, I210C, I210T, A211V, T229R, T229V, I230N, R233L, P2 34Q, P234V, T235H, D236R, D237G / H381W, E238L, E238R, E238W, H241R, H241S, V243M, V243S, V244S, L247 and an amino acid sequence having one or more residue differences or a set of residue differences selected from F, L253V, E260A, E260G, N278S, L286C, L286V, L288A, K292R, A297V, V298F, V298T, G299N, G299S, F327H, L331K, I368E, I368S, I368T, D373Q, D373R, and H381L.

[0215] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5628, and has at least one of the following sequences compared to SEQ ID NO: 5628: 26, 26 / 60 / 200, 26 / 60 / 200 / 203 / 292, 26 / 60 / 203 / 229 / 234 / 290, 26 / 200 / 203, 26 / 200 / 20 and 360. In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5628, and has at least one of the following sequences compared to SEQ ID NO: 5628: 26T, 26T / 60R / 200W, 26T / 60R / 200W / 203D / 292R, 26T / 60R / 203D / 229R / 234G / 290N, 26T / 200W / 203D, 26T / 200W / 203D / 229R, 26T / 2 and 360S.In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO:5628, and Compared to the following: I26T, I26T / K60R / R200W, I26T / K60R / R200W / S203D / K292R, I26T / K60R / S203D / T229R / P234G / R290N, I26T / R200W / S203D, I26T / R200W / S203D / T229R, I26T / R200W / R290N, I26T / S203D / P234G / R290N / K292R, K60R, K60R / R200W / S203D / R290N, K60R / S203D, A157V / S162F, A157V / I368Q, S162F / K242N / I368T, R200W, R200W / S203D / T229R / P234G, R200W / and amino acid sequences having one or more residue differences or sets of residue differences selected from S203D / R290N, R200W / A206P, R200W / P234G / R290N, R200W / R290N / K292R / G360S, S203D, S203D / T229R, T229R / G360S, and G360S.

[0216] In some embodiments, the engineered TdT polypeptide has at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the reference sequence SEQ ID NO: 5628, and has at least one of the following sequences compared to SEQ ID NO: 5628: 26, 26 / 60 / 200, 26 / 60 / 200 / 203 / 229 / 234 / 267, 26 / 60 / 200 / 203 / 229 ... 203 / 229 / 267 / 290, 26 / 60 / 200 / 203 / 234 / 267, 26 / 60 / 200 / 203 / 292, 26 / 60 / 200 / 234 / 290, 26 / 60 / 203 / 229 / 234 / 290, 26 / 60 / 229 / 267, 26 / 60 / 290, 26 / 200, 26 / 200 / 203, 26 / 200 / 203 / 229, 26 / 200 / 290, 26 / 229 / 234 / 360, 53 / 157 / 278 / 327 / 331, 53 / 162 / 327 / 331 / 368, 60, 60 / 20 0 / 203 / 290, 60 / 200 / 229 / 234, 60 / 200 / 234, 60 / 200 / 234 / 267 / 290 / 292, 60 / 200 / 234 / 290, 60 / 203, 157 / 162, 157 / 162 / 241 / 242 / 260, 157 / 162 / 241 / 242 / 327 / 331, 157 / 162 / 241 / 278 / 331 / 368, 157 / 162 / 241 / 331, 157 / 162 / 242 / 260 / 331, 157 / 162 / 331, 157 / 327 / 331, 162 / 241 / and amino acid sequences having one or more residue differences or sets of residue differences selected from 278 / 327 / 331, 162 / 241 / 331, 162 / 242 / 368, 200, 200 / 203 / 229 / 234, 200 / 203 / 290, 200 / 206, 200 / 234 / 290, 200 / 290 / 292 / 360, 210 / 211 / 242, 229 / 360, 241 / 242 / 368, 242 / 331, 260 / 327, 260 / 331, 260 / 368, 331, 360, and 368. In some embodiments, the engineered TdT polypeptide has a sequence identity that is at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, 110%, 111%, 112%, 113%, 114%, 115%, 116%, 117%, 118%, 119%, 120%, 121%, 122%, 123%, 124%, 125%, 126%, 127%, 128%, 129%, 130%, 131%, 132%, 133%, 134%, 135%, 136%, 137%, 138%, 139%, 140%, 141%, 142%, 143%, 144%, 145%, 146%, 147%, 148%, 149%, 150%, 151%, 152%, 153%, 154%, 155%, 156%, 157%, 158%, 159%, 160%, 161%, 162%, 163%, 164%, 165%, 166%, 167%, 168%, 169%, 170%, 171%, 172%, 173%, 174%, 175%, 176%, 177%, 178%, 179%, 180%, 181%, 182%, 183%,95%, 96%, 97%, 98%, 99% or higher sequence identity, and the following compared to SEQ ID NO: 5628: 26T, 26T / 60R / 200W, 26T / 60R / 200W / 203D / 229R / 234G / 267K, 26T / 60R / 200W / 203D / 229R / 267K, 26T / 60R / 200W / 203D / 229R / 267K / 290N, 26T / 60R / 200W / 203D / 292R, 26T / 60R / 200W / 203D / 234G / 267K, 26T / 60R / 200W / 203D / 292R, 26T / 60R / 200W / 234G / 290N, 26T / 60R / 200W / 203D / 292R ...34G / 290N, 26T / 60R / 200W / 203D / 292R, 26T / 60R / 200W / 234G / 290N, 26T / 60R / 200W 03D / 229R / 234G / 290N, 26T / 60R / 229R / 267K, 26T / 60R / 290N, 26T / 200W, 26 T / 200W / 203D, 26T / 200W / 203D / 229R, 26T / 200W / 290N, 26T / 229R / 234G / 36 0S, 53T / 157V / 278G / 327H / 331K, 53T / 162F / 327H / 331N / 368Q, 60R, 60R / 20 0W / 203D / 290N, 60R / 200W / 229R / 234G, 60R / 200W / 234G, 60R / 200W / 234G / 26 7K / 290N / 292R, 60R / 200W / 234G / 290N, 60R / 203D, 157L / 162F / 241S / 331K, 157V / 162F, 157V / 162F / 241S / 242N / 260A, 157V / 162F / 241S / 242N / 327H / 3 31K, 157V / 162F / 241S / 278G / 331K / 368Q, 157V / 162F / 242N / 260A / 331K, 15 7V / 162F / 331K, 157V / 327H / 331K, 162F / 241S / 278G / 327H / 331K, 162F / 241S In some embodiments, the amino acid sequence has one or more residue differences or a set of residue differences selected from: 210T / 211V / 242N, 229R / 360S, 241S / 242N / 368Q, 242N / 331K, 260A / 327H, 260A / 331K, 260A / 368Q, 331K, 360S, and 368Q.Engineered TdT polypeptides have at least 60%, 70%, 80%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or higher sequence identity to the reference sequence SEQ ID NO: 5628, and may have any of the following sequences compared to SEQ ID NO: 5628: I26T, I26T / K60R / R200W, I26T / K60R / R200W / S203D / T229R / P234G / Q267K, I26T / K60R / R200W / S203D / T229R / Q2...

Claims

1. SEQ ID NOs: 2, 4, 580, 692, 882, 914, 1034, 1270, 1344, 1346, 1678, 1700, 1750, 1932, 2164, 2666, 2794, 2978, 3074, 3302, 3398, 3488, 3958, 3788, 4124, 4226, 4734, 5052, 5152, 5252, 5296, 5628, 5630, 5632, and and / or an engineered terminal deoxynucleotidyl transferase comprising a polypeptide sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to a reference sequence of 5636, or a functional fragment thereof, wherein said engineered terminal deoxynucleotidyl transferase is a terminal deoxynucleotidyl transferase.

10. An engineered terminal deoxynucleotidyl transferase having nucleotidyl transferase activity, wherein the engineered terminal deoxynucleotidyl transferase comprises at least one residue difference or set of residue differences in its polypeptide sequence, wherein the amino acid positions of the polypeptide sequence are numbered with reference to SEQ ID NOs: 2, 4, 580, 692, 882, 914, 1034, 1270, 1344, 1346, 1678, 1700, 1750, 1932, 2164, 2666, 2794, 2978, 3074, 3302, 3398, 3488, 3958, 3788, 4124, 4226, 4734, 5052, 5152, 5252, 5296, 5628, 5630, 5632, and / or 5636.

2. 2. The engineered terminal deoxynucleotidyl transferase of claim 1, wherein the at least one residue difference or set of residue differences comprises a residue difference of C68R, wherein the amino acid positions are numbered with reference to SEQ ID NO: 5296.

3. 26, 26 / 60 / 200, 26 / 60 / 200 / 203 / 292, 26 / 60 / 203 / 229 / 234 / 290, 26 / 200 / 203, 26 / 200 / 203 / 229, 26 / 200 / 290, 26 / 203 / 234 / 290 / 292, 60, 60 / 200 / 203 / 290, 60 / 203, 157 / 162, 157 / 368, 162 / 242 / 368, 200, 200 / 203 3. The engineered terminal deoxynucleotidyl transferase of claim 2, further comprising a residue difference or set of residue differences at an amino acid position selected from: 200 / 229 / 234, 200 / 203 / 290, 200 / 206, 200 / 234 / 290, 200 / 290 / 292 / 360, 203, 203 / 229, 229 / 360, and 360, and / or any combination thereof, wherein the amino acid positions are numbered with reference to SEQ ID NO:5628.

4. 26, 26 / 60 / 200, 26 / 60 / 200 / 203 / 229 / 234 / 267, 26 / 60 / 200 / 203 / 229 / 267, 26 / 60 / 200 / 203 / 229 / 267 / 290, 26 / 60 / 200 / 203 / 234 / 267, 26 / 60 / 200 / 203 / 292, 26 / 60 / 200 / 234 / 290, 26 / 60 / 203 / 229 / 234 / 290, 26 / 60 / 229 / 267, 26 / 60 / 290, 26 / 200, 26 / 200 / 203 , 26 / 200 / 203 / 229, 26 / 200 / 290, 26 / 229 / 234 / 360, 53 / 157 / 278 / 327 / 331, 53 / 162 / 327 / 331 / 368, 60, 60 / 200 / 203 / 290, 60 / 200 / 229 / 234, 60 / 200 / 234, 60 / 200 / 234 / 267 / 290 / 292, 60 / 200 / 234 / 290, 60 / 203, 157 / 162, 157 / 162 / 241 / 242 / 260, 157 / 162 / 241 / 242 / 327 / 331, 157 / 162 / 241 / 278 / 331 / 368, 157 / 162 / 241 / 331, 157 / 162 / 242 / 260 / 331, 157 / 162 / 331, 157 / 327 / 331, 162 / 241 / 278 / 327 / 331, 162 / 241 / 331, 162 / 242 / 368, 200, 200 / 203 / 229 / 234, 200 / 203 / 290, 200 / 206, 200 / 234 / 290, 200 / 290 / 292 / 360, 3. The engineered terminal deoxynucleotidyl transferase of claim 2, further comprising a residue difference or set of residue differences at amino acid positions selected from 210 / 211 / 242, 229 / 360, 241 / 242 / 368, 242 / 331, 260 / 327, 260 / 331, 260 / 368, 331, 360, and 368, and / or any combination thereof, wherein said amino acid positions are numbered with reference to SEQ ID NO:5628.

5. The engineered terminal deoxynucleotidyl transferase of claim 2, further comprising a residue difference or set of residue differences at amino acid positions selected from 217, 226, 239, 256, 264, 273, 275, 315, 325, 328, 339, 342, 342 / 363, 344, 345, 346, 350, 351, 352, 353, 361, 363, 364, 365, 366, 367, 371, 374, 376, 380, 383, and 388, and / or any combination thereof, wherein the amino acid positions are numbered with reference to SEQ ID NO:5628.

6. 113 / 355, 134, 169 / 390, 217, 226, 239, 256, 264, 265, 271, 275, 313, 314, 315, 321, 325, 328, 332, 339, 342, 342 / 363, 344, 345, 346, 348, 349, 350, 351, 352, 353, 354, 356, 360, 361, 362, 363, 364, 365, 366, 367, 369, 370, 37 3. The engineered terminal deoxynucleotidyl transferase of claim 2, further comprising a residue difference or set of residue differences at an amino acid position selected from 1, 374, 376, 377, 378, 380, 382, ​​383, 384, 385, 386, 388, and 391, and / or any combination thereof, wherein said amino acid positions are numbered with reference to SEQ ID NO:5628.

7. 184 / 188 / 200 / 203 / 211 / 242, 184 / 188 / 200 / 203 / 242 / 297, 184 / 188 / 200 / 203 / 290 / 297 / 368 , 184 / 188 / 203 / 290 / 297, 184 / 188 / 211 / 242 / 290 / 368, 184 / 189, 184 / 189 / 206 / 297 / 368, 18 4 / 189 / 297, 184 / 200, 184 / 206 / 242 / 290 / 297, 184 / 211, 184 / 242, 184 / 242 / 297 / 368, 184 / 290 / 297, 184 / 290 / 368, 188 / 211, 188 / 211 / 242 / 290 / 297 / 368, 192 / 193 / 211 / 242 / 297 / 368, 3. The engineered terminal deoxynucleotidyl transferase of claim 2, further comprising a residue difference or set of residue differences at an amino acid position selected from 192 / 193 / 290 / 297 / 368, 193 / 194, 194, 194 / 242, 194 / 242 / 290, 200, 200 / 203, 200 / 203 / 290 / 368, 203 / 206 / 368, 203 / 242 / 290 / 297 / 368, 203 / 297, 206 / 290, 211, 211 / 368, 242 / 290, 242 / 290 / 297, 297, and 368, and / or any combination thereof, wherein the amino acid positions are numbered with reference to SEQ ID NO:5630.

8. 184, 184 / 188 / 200 / 203 / 211 / 242, 184 / 188 / 200 / 203 / 242 / 297, 184 / 188 / 200 / 2 03 / 290 / 297 / 368, 184 / 188 / 203 / 290 / 297, 184 / 188 / 211 / 242 / 290 / 368, 184 / 189 , 184 / 189 / 206 / 297 / 368, 184 / 200, 184 / 242, 184 / 242 / 297 / 368, 184 / 290 / 297, 184 / 290 / 368, 188 / 211 / 242 / 290 / 297 / 368, 189 / 200, 192 / 193 / 211 / 242 / 297 / 36 3. The engineered terminal deoxynucleotidyl transferase of claim 2, further comprising a residue difference or set of residue differences at an amino acid position selected from: 8, 192 / 193 / 290 / 297 / 368, 194 / 242, 200, 200 / 203, 200 / 203 / 290 / 368, 203 / 206 / 368, 203 / 297, 206 / 290, 206 / 297, 211 / 368, 242 / 290, 242 / 290 / 297, 297, and 368, and / or any combination thereof, wherein the amino acid positions are numbered with reference to SEQ ID NO:5630.

9. The engineered terminal deoxynucleotidyl transferase of claim 2, further comprising a residue difference or set of residue differences at amino acid positions selected from 56, 57, 59, 63, 65, 66, 73, 128 / 296, 140, 144 / 220, 149, 151, 159, 209 / 211, 210 / 211, 219, 219 / 300, 230, 261, 272, 273, 275, 278, 292, 324, 327, 331, 345, 364, 371, and 373, and / or any combination thereof, wherein the amino acid positions are numbered with reference to SEQ ID NO:5632.

10. 56, 57, 59, 61, 65, 66, 73, 74 / 272, 75, 79, 128, 128 / 296, 134, 140, 144, 145, 149, 151, 155, 156, 157, 158, 159, 200 / 202 / 203, 209 / 211, 210 / 211, 211, 219, 219 / 300, 228, 230, 242, 242 / 244, 261, 262, 270, 272, 273, 275, 278 3. The engineered terminal deoxynucleotidyl transferase of claim 2, further comprising a residue difference or set of residue differences at an amino acid position selected from the following: 292, 319, 322, 324, 327, 339, 344, 345, 364, 371, 373, and 381, and / or any combination thereof, wherein the amino acid positions are numbered with reference to SEQ ID NO:5632.

11. 65, 65 / 70 / 155 / 209 / 228, 65 / 128 / 209 / 371 / 373, 65 / 151, 65 / 155 / 209, 65 / 155 / 209 / 352 / 353, 65 / 159, 65 / 209 / 219 / 352 / 353, 65 / 209 / 352 / 353 / 371, 65 / 228 / 352 / 353 / 373, 65 / 228 / 371 / 373, 65 / 352 / 353 / 371, 128 / 159 / 209 / 352 / 353, 128 / 20 9, 128 / 209 / 219, 128 / 209 / 219 / 352 / 353 / 371, 128 / 209 / 228 / 371, 128 / 209 / 371, 128 / 209 / 371 / 373, 128 / 209 / 373, 128 / 338 / 352 / 353 / 373, 151 / 209, 151 / 209 / 371, 151 / 209 / 373, 151 / 228 / 352 / 353 / 373, 151 / 228 / 373, 151 / 352 / 353, 151 / 352 / 353 / 371 / 373, 155, 155 / 209, 155 / 228 / 352 / 353, 159 / 209 / 352 / 353, 208 / 209, 209, 209 / 219 / 352 / 353 / 371, 209 / 219 / 371, 209 / 228, 209 / 352 / 353 / 371, 209 / 371, 209 / 371 / 373, 209 / 373, 219 / 228, 219 / 352 / 353 / 371 / 373, 219 / 371 / 373, 228, 3. The engineered terminal deoxynucleotidyl transferase of claim 2, further comprising a residue difference or set of residue differences at amino acid positions selected from 228 / 371, 228 / 371 / 373, 228 / 373, 352 / 353, 352 / 353 / 371, 371, 371 / 373, and 373, and / or any combination thereof, wherein said amino acid positions are numbered with reference to SEQ ID NO:5636. Request 12: 55 / 65 / 151, 65, 65 / 70 / 155 / 209 / 228, 65 / 70 / 228 / 373, 65 / 128, 65 / 128 / 155 / 209 / 371, 65 / 128 / 159 / 209 / 371 / 373, 65 / 128 / 209 / 371, 65 / 128 / 209 / 371 / 373, 65 / 128 / 219 / 373, 65 / 128 / 373, 65 / 151, 65 / 155 / 209, 65 / 155 / 209 / 352 / 353, 65 / 159, 65 / 209 / 219 / 352 / 353, 65 / 209 / 352 / 353 / 371, 65 / 219 / 371, 6 5 / 228 / 352 / 353 / 373、65 / 228 / 371、65 / 228 / 371 / 373、65 / 352 / 353、65 / 352 / 353 / 371、65 / 373、70 / 155 / 352 / 353、128、128 / 155 / 209、128 / 159、128 / 159 / 209 / 352 / 353、128 / 209、128 / 209 / 219、128 / 209 / 219 / 352 / 353 / 371、128 / 209 / 228 / 371、128 / 209 / 371、128 / 209 / 371 / 373、128 / 209 / 373、128 / 219 / 352 / 353、128 / 219 / 371 / 373、128 / 228 / 352 / 353、128 / 228 / 371、128 / 228 / 373、128 / 352 / 353、128 / 352 / 353 / 373、128 / 371、128 / 371 / 373、128 / 373、151、151 / 155、151 / 155 / 209、151 / 155 / 209 / 219 / 228 / 371 / 373、151 / 155 / 209 / 373、151 / 155 / 219 / 352 / 353 / 371、151 / 155 / 352 / 353、151 / 155 / 352 / 353 / 371 / 373、 151 / 155 / 352 / 353 / 373、151 / 209、151 / 209 / 371、151 / 209 / 371 / 373、151 / 209 / 373、151 / 219、151 / 219 / 371、151 / 228 / 352 / 353 / 373、151 / 228 / 373、151 / 352 / 353, 151 / 352 / 353 / 371 / 373, 151 / 371, 151 / 373, 155, 155 / 209, 155 / 219, 155 / 228 / 352 / 353, 155 / 228 / 371, 155 / 228 / 371 / 373, 155 / 352 / 353 / 373,155 / 371, 155 / 371 / 373, 155 / 373, 159 / 209 / 352 / 353, 209, 209 / 219 / 352 / 353 / 371, 209 / 219 / 371, 209 / 228, 209 / 352 / 353 / 371, 209 / 371, 209 / 371 / 373, 209 / 373, 219, 219 / 228, 219 / 352 / 353 / 371 / 373, 219 / 352 / 353 / 373, 219 / 371, 219 / 371 / 373, 228 / 352 / 35 3, 228 / 371, 228 / 371 / 373, 228 / 373, 352 / 353, 352 / 353 / 371, 352 / 353 / 373, 371, 371 / 373, and 373, and / or any combination thereof, wherein the amino acid positions are numbered with reference to SEQ ID NO:5636.

13. 1. An engineered terminal deoxynucleotidyl transferase comprising a polypeptide sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to a reference sequence of SEQ ID NOs:5296-6766, or a functional fragment thereof, wherein the engineered terminal deoxynucleotidyl transferase comprises at least one residue difference or set of residue differences in the polypeptide sequence, and wherein the amino acid positions of the polypeptide sequence are numbered with reference to SEQ ID NOs:5296, 5628, 5630, 5632, and / or 5636.

14. An engineered terminal deoxynucleotidyl transferase comprising a polypeptide sequence selected from the even-numbered sequences of SEQ ID NOs: 5296-6766.

15. The at least one residue difference or set of residue differences is 8, 9, 10, 11, 12, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 26, 27, 28, 29, 30, 32, 34, 36, 38, 40, 42, 44, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 69, 70, 71, 72, 73, 74, 75, 77, 78, 79, 80, 82, 84, 85, 86, 87, 89, 91, 92, 93, 94, 96, 97, 98, 99, 100 , 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 115, 116, 117, 118, 119, 120, 122, 123, 124, 125, 127, 131, 133, 134, 135, 136, 140, 141, 143, 144, 145, 147, 148, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 170, 173, 174, 175, 176, 177, 180, 181, 182, 184, 1 85, 186, 187, 189, 190, 192, 193, 194, 196, 197, 198, 200, 201, 202, 203, 204, 205, 206, 207, 209, 210, 211, 217, 219, 220, 224, 226, 228, 229, 230, 231, 2 32, 233, 234, 235, 236, 237, 238, 239, 241, 242, 243, 244, 245, 246, 248, 249, 250, 252, 253, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 26 8, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 282, 284, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302 , 303, 304, 305, 306, 307, 308, 309, 310, 311, 315, 319, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 332, 338, 339, 340, 341, 342, 344, 345, 346, 347,2. The engineered terminal deoxynucleotidyl transferase of claim 1, further comprising a residue difference or set of residue differences at an amino acid position selected from 349, 350, 351, 352, 353, 355, 358, 360, 361, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 376, 377, 378, 380, 383, 384, 386, 387, 388, and 390, and / or any combination thereof, wherein said amino acid positions of said polypeptide sequence are numbered with reference to SEQ ID NOs: 5296, 5628, 5630, 5632, and / or 5636.

16. The at least one residue difference or set of residue differences is 8, 9, 9 / 11, 9 / 14 / 190 / 197 / 364, 9 / 14 / 58, 9 / 14 / 58 / 182 / 190 / 197 / 298, 9 / 28 / 156 / 158 / 173 / 190 / 193 / 303 / 364, 9 / 28 / 156 / 158 / 290 / 364, 9 / 28 / 190 / 193 / 303 / 364, 9 / 28 / 290 / 303 / 364, 9 / 58 / 197, 9 / 156 / 158 / 193 / 290 / 303 / 364, 9 / 156 / 364, 9 / 290 / 364, 9 / 298, 9 / 302, 9 / 3 64, 10, 10 / 14 / 197 / 364, 11, 12, 14, 14 / 15 / 200 / 230, 14 / 15 / 77 / 200 / 226 / 230, 14 / 15 / 77 / 78, 14 / 15 / 77 / 78 / 200 / 201 / 226 / 230 / 290, 14 / 15 / 77 / 78 / 200 / 203 / 230 / 290, 14 / 15 / 77 / 78 / 203 / 226 / 230 / 290, 14 / 15 / 78, 14 / 15 / 78 / 226 / 230, 14 / 28 / 224 / 296 / 298, 14 / 28 / 58 / 162 / 190 / 193 / 224, 14 / 28 / 58 / 224, 1 4 / 28 / 58 / 84 / 173 / 297, 14 / 58 / 147 / 162 / 190 / 193 / 224 / 296 / 303, 14 / 58 / 158 / 224, 14 / 58 / 162, 14 / 58 / 182 / 197 / 298 / 364, 14 / 58 / 84 / 147 / 162 / 224 / 296 / 297 / 298 / 299, 14 / 58 / 84 / 147 / 162 / 224 / 296 / 297 / 299, 14 / 58 / 84 / 224 / 298, 14 / 77 / 201 / 230, 14 / 77 / 78 / 200 / 226 / 230 / 257, 14 / 77 / 78 / 226 / 230 / 257, 14 / 78 / 200 / 201 / 226 / 290, 14 / 78 / 200 / 201 / 339, 14 / 78 / 200 / 226 / 230 / 339, 14 / 78 / 200 / 226 / 290 / 339, 14 / 78 / 200 / 230 / 339, 14 / 84 / 147 / 193 / 197 / 224 / 296, 14 / 84 / 162 / 224 / 297 / 299, 14 / 84 / 224 / 296 / 299, 14 / 147, 14 / 147 / 158 / 162 / 224, 14 / 147 / 173, 14 / 147 / 224, 14 / 158 / 162 / 197 / 224 / 296 / 298 / 299,１４／１６２／２２４／２９８／２９９、１４／１７３／１９３／１９７／２９６／２９７／２９８／２９９、１４／１９０／１９７／２９８、１４／１９３／１９７／２２４／２９７／２９８／２９９／３００、１４／１９７／２９８／３６４、１４／２００／２０３／２２６／２３０、１４／２００／２０３／２２６／２３０／２５７／３３９、１４／２００／２０３／２３０／３３９、１４／２０１／２０３／２２６、１４／２２４／２９６／２９８、１４／２２４／２９６／３０３、１４／２９８／３６４、１５、１５／７７、１５／７７／２００／２０１／２０３／２２６／２３０／２９０／３３９、１５／７７／２００／２３０、１５／７７／２００／２３０／２９０／３３９、１５／７７／２０１／２２６／２３０／３３９、１５／７７／２２６／２３０／２９０／３３９、１５／７７／７８、１５／７７／７８／２００／２０１／２０３／２２６／２３０、１５／７８／２０１、１５／７８／２０１／２０３／２３０、１５／７８／２０１／２３０／２９０、１５／７８／２０３／２２６／２３０、１５／２００、１５／２０１／２０３／２２６／２３０／２５７／３３９、１５／２２６／２３０／３３９、１５／２３０、１６、１７、１８、１８／２８／１４７／３０３、１９、２０、２０／２１、２０／２１／１０３／２３３、２０／２１／１１１、２０／２１／１１１／１５７／３１５、２０／２１／１１１／１６０／２００、２０／２１／１１１／２００／２３８、２０／２１／１１１／２４６、２０／２１／１８０／２９７、２０／２１／２３３、２０／２１／３１５、２０／２１／５２／６７／６８／８７／１１１／１５７／１６０／１７３／１８０／２００／２３５／２４６／３１５、２０／２１／５５／６７／６８／８７／１１１／１５７／１６０／１８０／１８１／２００／２３５／２４６／２５６／３１５／３５０、２０／２１／５５／６７／６８／８７／１１１／１５７／１６０／１８０／２００／２４６／２５６／３１５、２０／２１／６０／７２／１６０／１８０／２００／２４６／２５９／３３８／３５８、２０／２１／６７、２０／２１／６７／１６０、２０／２１／６７／６８／８７／１１１／１５７／１６０／１８０／２００／２３５／２４６／３１５、２０／２１／６７／６８／８７／１１１／１５７／１６０／１８０／２００／２４６／３１５、２０／２１／６７／８７、２０／２１／６７／８７／１１１、２０／２１／６７／８７／１１１／２００、２０／２１／６７／８７／１１１／３１５、２０／２１／６７／８７／１５７／１８０、２０／２１／６７／８７／２４６／３１５、２０／２１／６８／１０３／２００／２３８／２９７、２０／２１／６８／１１１／２３５、２０／２１／６８／１６０、２０／２１／６８／１６０／１８０／２００／２４６、２０／２１／６８／１６０／２４６、２０／２１／６８／１８０、２０／２１／６８／１８０／２３５、２０／２１／６８／２００／２３５／２９７、２０／２１／６８／２３３／２４６／２９７、２０／２１／８７／１１１／２４６／３１５、２０／２１／８７／１５７／２００、２０／２１／８７／１８０／２４６／３１５、２０／２１／８７／３１５、２０／６７／８７／１１１／１８０、２０／６７／８７／１５７／１６０／１８０／３１５、２０／６８／１０３／１６０／２００、２０／６８／１０３／１８０／２００、２０／６８／１１１／１５７／１６０、２０／６８／１６０、２０／６８／１６０／１８０、２０／６８／１６０／１８０／２３３／２３５／２４６、２０／６８／１６０／２００／２４６、２０／６８／１６０／２３５／２９７、２０／６８／１６０／２４６、２０／６８／２００／２３８／２４６、２０／６８／２３５／２９７、２０／８７／１１１／１８０、２０／８７／１５７／１６０、２０／１００、２０／１００／１０４／１１１／２４２、２０／１００／１０４／１２０／１９７／３６７、２０／１００／１０４／１９７／２０３／２４２／２９２、２０／１００／１０４／２０３／３５０／３５３、２０／１００／１１１、２０／１００／１２０／１９７／２３２／２３５／３１５／３６６／３６７、２０／１００／１９７／２９２／３１５、２０／１００／２０３、２０／１００／２３２／２９２／３６６／３６７、２０／１００／２３５／３１５／３６７、２０／１０３／１６０／１８０／２００／２３５、２０／１０３／２３３、２０／１０４／１１１／１２０／２０３、２０／１１１、２０／１１１／１８０／２３５／２４６／２９７、２０／１２０／１９７／２９２、２０／１２０／２３５、２０／１６０／３１５、２０／２００、２０／２９２、２１、２１／６７／１８０／３１５、２１／６７／２００、２１／６７／８７／１１１／２００、２１／６７／８７／１５７／２４６、２１／６８、２１／６８／１０３／１１１、２１／６８／１１１／２００、２１／６８／１６０／１８０／２００／２０５／２９７、２１／６８／１６０／１８０／２００／２９７、２１／６８／１６０／２３８、２１／６８／１６０／２３８／２４６、２１／６８／１８０、２１／６８／１８０／２３５、２１／６８／１８０／２４６、２１／６８／２００、２１／６８／２３５、２１／８７／１５７／１６０、２１／８７／１６０／２００／３１５、２１／８７／１６０／３１５、２１／８７／２００、２１／８７／２４６／３１５、２１／１０３／２３３、２１／１１１、２１／１５７／１６０、２１／１６０／３１５、２１／２００／３１５、２１／２３３／２９７、２１／２４６、２２、２３、２３／２８／１５６／１５８／１９０／１９３／２９０／３６４、２３／１８６／２５６／３０９、２４、２６、２６／６０／２００、２６／６０／２００／２０３／２２９／２３４／２６７、２６／６０／２００／２０３／２２９／２６７、２６／６０／２００／２０３／２２９／２６７／２９０、２６／６０／２００／２０３／２３４／２６７、２６／６０／２００／２０３／２９２、２６／６０／２００／２３４／２９０、２６／６０／２０３／２２９／２３４／２９０、２６／６０／２２９／２６７、２６／６０／２９０２６／２００、２６／２００／２０３、２６／２００／２０３／２２９、２６／２００／２９０、２６／２０３／２３４／２９０／２９２、２６／２２９／２３４／３６０、２７、２８、２９、２９／７７／１０４／２３４／２７１、２９／７７／１０４／２３４／２７１／２７９／３８０、２９／７７／１８２／２０７／２７１、２９／７７／２０７／２３４／２７９／３８０、２９／７７／２３４／２７１／２７９／３６８、２９／７７／２７１、２９／９８／１４３／２６６、２９／９８／１８５／１９７／２６６、２９／９８／１８５／２６６／２９６／２９９、２９／１０４／２３４／２７１／２７９／３８０、２９／１４３、２９／１４３／１６２、２９／１４３／１７０／１９３／１９７／２６６、２９／１４３／１８５／１９３／２６６、２９／１５８／２６６、２９／１７０／１８５／１９３／２６６／２９９、２９／１８２／２０７／３８０、２９／１８２／２３４／２７１／３８０、２９／１８２／２７１／２７９／３８０、２９／１８５／２９６、２９／１９３／１９７／２９６／２９７、２９／２３４／２７９／３８０、２９／２７１／２７９、２９／２７１／２７９／３６８／３８０、３０、３２、３２／７２／３３９、３２／７２／３３９／３６０、３２／７８／１０６／２００／２２６／２７２／３７３、３２／７８／１０６／２００／２２６／３７３、３２／７８／１０６／２２６、３２／７８／２００／２２６、３２／７８／２００／２２６／２３５／２７２、３２／７８／２００／２２６／２３５／３２１／３７３、３２／７８／２２６、３２／７８／２２６／３２１、３２／７８／２２６／３７３、３２／１０３、３２／１０３／１０６／１１１／３２２／３２４、３２／１０３／１０６／２０７／２１０／２３５／３２１／３６８、３２／１０３／２０７／２１０／３４４／３４９、３２／１０３／２０７／２２８／２３２／３７３、３２／１０３／２０７／３２１／３４４、３２／１０３／２１０／２７３／３２１／３７３、３２／１０３／２７３、３２／１０３／３２１／３４４、３２／１０６、３２／１０６／２００／２２６、３２／１０６／２００／２２６／２３５／３７３、３２／１０６／２０７／２１０／２３２、３２／１０６／２１０、３２／１０６／２１０／３７３、３２／１０６／２３５／３２１、３２／１０６／２３５／３６８、３２／１０６／２７３／３２１／３７３、３２／１０６／２８０、３２／１１１／２３５、３２／１１１／２３５／２７１／２７２／３３９、３２／１１１／２３５／３３９、３２／１１１／２３５／３３９／３８６、３２／１１１／２３５／３８６、３２／１４０／１４１／２６１／３６０、３２／１４０／３３９、３２／１４０／３６０、３２／１４１／１８０／２４４、３２／１６１／１９３／２３７／３６０、３２／１６２、３２／１６２／１９３、３２／１６２／１９３／２６７、３２／１６２／２３７／２６５／２６６、３２／１６２／３０２、３２／１８０／２６１／３３９、３２／２００／２２６、３２／２００／２２６／３２１／３７３、３２／２０７／２１０／２７３、３２／２０７／２１０／２７９、３２／２０７／２１０／３６８／３７３、３２／２０７／２３５、３２／２０７／２７３／２７９／３２１、３２／２０７／２７９、３２／２０７／３４４、３２／２１０／２３２／２３５／３６８、３２／２１０／２３２／２７３／３２１、３２／２１０／２３２／２７３／３６８／３７３、３２／２１０／２７３、３２／２２６、３２／２３５、３２／２３５／２７１／３３９、３２／２３５／２７２、３２／２３５／２７２／３３９、３２／２３５／２７２／３８６、３２／２３５／２７３、３２／２３５／３３９／３８６、３２／２３５／３８６、３２／２４、 ４／２６１、３２／２７１／３３９、３２／２７２、３２／２７３／２７９／３４４／３４９、３２／２７３／３４４／３７３、３２／２７９、３２／２７９／３２１、３２／３０２、３２／３２１／３２４／３６０、３２／３２２／３２４／３８３／３８６、３２／３２４、３２／３３９、３４、３４／４８／１３３／１５８／１８２／２３０／２３３／２７１／３４５、３４／４８／１４７、３４／４８／１４７／１５８／１８２／２２０／２３３／３０７、３４／４８／１４７／１８２／２３０／２３３／２４９／３０７、３４／４８／１４７／２７１、３４／４８／１８２／２３３／２４９、３４／４８／３３９、３４／４８／７８／１３３／１４７／１８２、３４／４８／７８／１４７／１５８、３４／４８／７８／１４７／１８２／２２０／２３３／２４９／３０７／３１５／３３９、３４／４８／７８／１５８／１８２／２２０／２４９／３０７、３４／４８／７８／１５８／１８２／２３３／３１５／３４５、３４／４８／７８／１８２／２２０／２３０／３１５、３４／４８／７８／１８２／２２０／２３３、３４／７８、３４／７８／１４７／１８２／２２０／２４９、３４／７８／１４７／１８２／２３３／２４９／３１５、３４／７８／１４７／１８２／２３３／２７１／３３９、３４／７８／１５８、３４／７８／１５８／１８２／３１５／３４５、３４／７８／１５８／２４９、３４／７８／１８２／２３３／３０７、３４／７８／２０４／２２０／３３９、３４／７８／２２０／２７１、３４／７８／２２０／３０７／３３９、３４／７８／３１５、３４／７８／８４／１４７／１５８／１８２／２２０、３４／７８／８４／１５８／２３０、３４／１３３／１４７／１５８／２３０／２３３／２４９、３４／１４７、３４／１４７／１５５／２３３／３３９、３４／１４７／１５８、３４／１４７／１５８／１８２／２３３、３４／１４７／１５８／１８２／２４９／２７１、３４／１４７／１８２、３４／１４７／１８２／２２０、３４／１４７／１８２／２２０／２３０／２４９／３１５／３３９、３４／１４７／１８２／２２０／２３０／３３９、３４／１４７／１８２／２２０／２７１／３１５、３４／１４７／１８２／２３３／２７１／３３９、３４／１４７／１８２／２４９／３０７／３１５／３３９、３４／１４７／１８２／３１５、３４／１４７／１８２／３４５、３４／１４７／２２０／２７１、３４／１４７／２３０／２７３／３１５／３４５、３４／１４７／２３３、３４／１４７／２４９、３４／１４７／３１５／３３９、３４／１５８／１８２／３１５／３３９／３４５、３４／１５８／２２０、３４／１５８／３０７、３４／１８２、３４／１８２／２３０／３１５、３４／１８２／３０７／３３９、３４／１８２／３４５、３４／２２０、３４／２２０／３０７、３４／２２０／３０７／３４５、３４／２２０／３１５／３３９、３４／２２０／３３９、３４／２３３、３４／２７１／３３９、３４／３１５／３４５、３４／３３９、３６、３８、４０、４２、４４、４４／１９３、４７、４８、４８／５３／２３７／２３９、４８／７８／１４７／１５８／１８２／２２０／２３０／３０７、４８／７８／１４７／１５８／１８２／２３０／２４９／２７１／３１５、４８／７８／１４７／１５８／２３３／２４９、４８／７８／１４７／１８２／２２０／２３３／２４９／３３９／３４５、４８／７８／１４７／１８２／２２０／３１５、４８／７８／１４７／１８２／２３０／２３３／２４９、４８／７８／１５８／１８２／２２０／２３３／２４９、４８／７８／１５８／２３０／３３９、４８／７８／２３３／３１５／３３９、４８／１４７／１５８／１８２／２２０／２３０／２４９／２７１／３０７／３１５／３３９、４８／１４７／１５８／１８２／２２０／２４９／３０７／３３９、４８／１４７／１５８／１８２／２２０／３１５、４８／１４７／１５８／１８２／２３０／２３３、４８／１４７／１５８／１８２／２３３／３４５、４８／１４７／１５８／２３３、４８／１４７／１５８／３０７／３４５、４８／１４７／２３３、４８／１４７／２３３／３４５、４８／１４７／２７１／３０７／３３９、４８／１５８／１８２／２３０／２３３／２４９、４８／１８２、４８／１８２／３０７／３１５、４８／２５６／２６１、４８／３３９、４９、５０、５１、５２、５２／５５／１０６／２５６、５２／５５／１８１／２３５／２５６、５２／５５／１８１／２５６、５２／９９／１８１／２３５、５２／１０６／１８１／２３５／２５６、５２／１０６／２３５／２５６、５２／１０６／２５６、５２／１７３／２３５、５２／１８０、５２／１８０／２００／２３５／３１５／３４９、５２／１８０／２００／３４９、５２／１８０／３４９、５２／２００、５２／２００／３１５、５２／２００／３４９、５２／２００／３４９／３５０、５２／２３５、５２／２３５／２５６、５２／３１５、５２／３１５／３４９、５２／３４９、５２／３４９／３５０、５３、５３／７３／７５／２３７／２３９、５３／１５７／２７８／３２７／３３１、５３／１６２／３２７／３３１／３６８、５３／１６３／２０１／３２５／３２９、５３／２００／２０１／３２５／３２９、５３／２０１、５３／２０１／２７５／２８０、５３／２０１／３７１、５３／２１９／３５８、５３／２３７、５４、５５、５５／５８、５５／５８／２５６、５５／５８／２５６／３５０／３７３、５５／５８／２５６／３５５、５５／５８／３５０／３５１、５５／５８／６９／３５０／３５１、５５／５８／９９／２５６／３５１／３７３、５５／５８／９９／２５６／３５５、５５／５８／９９／３５１、５５／６７、５５／６７／１０６／１１１／１５７、５５／６７／１０６／３１５、５５／６７／１１１、５５／６７／１１１／３１５、５５／６７／３１５、５５／６７／８７、５５／６７／８７／１０６／１１１／３１５、５５／６７／８７／１５７／３１５、５５／８０／１７４／２６８／３５５／３６６、５５／８０／２６８／３１５／３６６、５５／８０／２６８／３４６、５５／８７、５５／８７／１０６、５５／８７／１０６／１１１／３１５、５５／８７／１０６／３１５、５５／８７／１５７、５５／８７／１５７／２０７、５５／９９／１０３、５５／９９／１８１／２５６、５５／９９／２１９／３５８／３７３、５５／９９／２５６／３５０、５５／１０３／１８１、５５／１０３／２１９、５５／１０３／３３８、５５／１０６／１１１、５５／１０６／１５７、５５／１１１、５５／１１１／１５６／２６８／３１５／３２４／３２７／３６６／３７３、５５／１１１／２６８／３４６／３５５、５５／１１１／２６８／３５５／３６６、５５／１１１／３１５、５５／１１１／３１５／３５５／３７３、５５／１５７、５５／１８１／２１９、５５／１８１／２３５／２５６／３５０、５５／１８１／２４６、５５／１８１／３５８、５５／２１９／２４６／３５８、５５／２１９／２５６／３３８、５５／２５６、５５／２５６／２５９、５５／２６８、５５／２６８／３１５／３４６、５５／２６８／３２４／３６６、５５／２６８／３４６／３５５、５５／３１５、５６、５６／７５／１５４／１５６／１９２／２３９／２８０／２８２、５６／７５／１９２／２３９、５６／１９２／２８２／３５０、５７、５７／３６７、５８、５８／６９／２５６／３７３、５８／７２／２１１／３１５、５８／７２／２２０／２２４、５８／８４／２１１／２２０／２２４、５８／９９、５８／９９／３５１／３５５、５８／９９／３５５、５８／１４７／１６２／１９７／２２４／２９６／２９７／２９８／２９９、５８／１４７／１６２／２９６／２９８、５８／１６２／２２４／２９６／２９８、５８／１８６／２７０、５８／１９７、５８／１９７／３６４、５８／２２４、５８／２２４／２９９、５８／２５６、５８／２５６／３５０／３５５、５８／３５０／３５５、５８／３６４、５９、５９／６２／６３／６８、５９／６２／６３／６８／１０３／２３４、５９／６２／６３／６８／１４７、６０、６０／６２／６８／９１／１１１／２３４／２８９、６０／１０６／１１１／２３５／３６０、６０／２００／２０３／２９０、６０／２００／２２９／２３４、６０／２００／２３４、６０／２００／２３４／２６７／２９０／２９２、６０／２００／２３４／２９０、６０／２０３、６０／２５９、６０／２７８、６０／２８０／３６０、６１、６２、６２／６３／６８、６２／６３／６８／９１／１０９／２１０、６２／６３／６８／９１／１４７／２０５／２１０／２３４、６２／６６／１００／１０１／１０４／２０３／２３５／３３８、６２／６６／６９／１４３／３３８／３５３、６２／６８／１０３、６２／６８／１１１、６２／６８／９１／１１１／２８９、６２／６９／１００／２３５／２６８／３４６、６２／６９／３５３、６２／６９／８０／１０１／１０４／１４３／２３５／３３８、６２／１１１／２３５／３１５／３５５、６２／１１１／２３５／３２４／３４６、６２／２０３／２１１／２３５／３３８／３５０、６２／２３５／２６８／３２７／３４６／３５０／３５３／３５５、６２／２３５／３４６／３５０／３５５、６２／３１５／３２７／３５３、６２／３２３／３４６／３５３／３５５、６３、６４、６５、６５／７０／１５５／２０９／２２８、６５／１２８／２０９／３７１／３７３、６５／１４０、６５／１４０／１９２／１９３、６５／１４０／１９２／１９３／３０２、６５／１５１、６５／１５５／２０９、６５／１５５／２０９／３５２／３５３、６５／１５９、６５／１８４／１８７、６５／１９３、６５／２０９／２１９／３５２／３５３、６５／２０９／３５２／３５３／３７１、６５／２２０／３３９、６５／２２８／３５２／３５３／３７３、６５／２２８／３７１／３７３、６５／２５９、６５／３５２／３５３／３７１、６６、６６／６９／１４３／２３５／３３８、６６／１００／２３５／３１５／３２７／３５３／３５５、６６／１１１／３４６／３５３／３５５、６６／２２０／２２４、６６／２３５／２６８／３４６、６６／２３５／３４６、６６／２３５／３７３、６７、６７／８７、６７／８７／１０６／１５７、６７／８７／１１１／１５７／１６０／３１５、６７／８７／１１１／１５７／３１５、６７／８７／１５７、６７／８７／３１５、６７／１０６、６７／１０６／１１１、６７／１０６／１１１／３１５、６７／１０６／１５７、６７／１１１、６７／１１１／３１５、６７／１５７、６７／１５７／１６０／１８０、６７／１５７／１６０／３１５、６７／１５７／３１５、６７／１６０、６７／１８０／２００、６７／１８０／２００／３１５、６７／３１５、８７、１０３／１６０／２３５、１０３／２００／２３５／２４６／２９７、１０６／２００、１０６／３２１／３２２、１１１／２００／２３８、１１１／２３３／２３６／２９７、１１８、１５７／１６０／２００／３１５、１６０／２３３／２４６、２００、２００／２３５／２９７、２００／２７０／３２１、２７０／３２１／３２２、３４４、３４４／３８３、６９、６９／８０／２０３／２１１／２７８／３３８、６９／１００、６９／１００／１１１／２９８／３５３／３５５、６９／１００／２３５、６９／１００／３５３／３６６、６９／１１１／２３５、６９／１１１／２３５／３００／３５３／３５５、６９／１１１／２３５／３１５、６９／２２０、６９／２３５／３１５、６９／２３５／３５３、６９／２６８、６９／２６８／３２４／３２７／３５３／３５５、６９／２６８／３４６、６９／２６８／３４６／３５３、６９／３１５／３５３、６９／３２４／３２７／３４６、６９／３２４／３４６／３５０、６９／３２４／３５３、６９／３３９、６９／３５３／３５５、７０、７０／７１、７０／７１／３５３、７０／７２、７０／７２／１４０／２４４／２６１／３３９、７０／７２／１４１／２４４、７０／７２／１８０／３６０、７０／１３４／３５３、７０／１４０／１４１／３３９、７０／２６ １／３３９／３６０、７０／３５３、７０／３６０、７１、７１／７７／１３３／３５３、７１／７７／３５３、７１／３５３、７２、７２／７４／２００／２７２／３３９／３４７、７２／８４／２２０／２２４／３１５、７２／８４／８６／２２４、７２／８６／２２０、７２／１８０／２４４／３３９、７２／２２０／２２４、７２／２２０／３１５、７２／２５６／３６０、７２／３６０、７３、７４、７４／１０６／２７０／３４４、７４／２００／３３９、７４／２７２、７５、７５／２０７／３７３／３７８、７５／２３３／３４４、７５／２３３／３６６、７５／２３７／３５０、７７、７７／７８／２００／２２６／３３９、７７／７８／２０１／２２６／２５７／３３９、７７／７８／２０３、７７／７８／２０３／２３０／３３９、７７／１３３／３５３、７７／１３４／３５３、７７／１８２／２７９／３８０、７７／１８２／３６８、７７／２００／２０１、７７／２００／２０１／２２６／２３０／３３９、７７／２００／２０３／２３０／２５７／３３９、７７／２００／２３０、７７／２３４／２７１／２７９／３８０、７７／３５３、７８、７８／１０６、７８／１０６／２２６／３２１／３７３、７８／１０６／２３５／３２１／３７３、７８／１２７、７８／１３３／１４７／１５８／１８２／２２０／２７１／３３９、７８／１３５／１８２／２３３／２４９／３１５／３４５、７８／１４７／１５８／１８２、７８／１４７／１５８／１８２／２３０／２４９、７８／１４７／１５８／１８２／２３３／２７１／３０７／３４５、７８／１４７／１５８／１８２／３３９／３４５、７８／１４７／１５８／２２０／２３０／２３３／２４９／２７１／３０７／３１５／３４５、７８／１４７／１５８／２４９／３０７／３１５、７８／１４７／１８２／２３０／２３３／２４９、７８／１４７／１８２／２４９／３０７、７８／１４７／１８２／３３９／３４５、７８／１４７／２３０／３０７、７８／１４７／２３３、７８／１４７／２４９／２７１／３３９、７８／１５８／１８２、７８／１５８／１８２／２３３／２７１／３１５、７８／１５８／１８２／３０７／３１５／３４５、７８／１８２／２２０／３３９、７８／１８２／２７１／３１５／３３９、７８／２００／２２６／３２１／３７３、７８／２００／２３０、７８／２０３／２３０／２９０、７８／２２６／２３０、７８／２２６／２９０、７８／２２６／３２１、７８／２３３、７８／３３９、７９、８０、８０／１１１／２６８／３２４／３２７／３４６／３６６／３７３、８０／１１１／３５５／３６６、８０／１４３／２０３／２１１／３３８、８０／２６８／３１５／３４６／３５５、８０／２６８／３２７／３４６／３６６、８０／２６８／３４６、８０／３１５／３４６／３６４／３７３、８０／３４６／３６６、８２、８２／１５４／２９６、８２／１８４／２２０／３８６、８４、８４／９２／１７３、８４／１４７、８４／１４７／１９７／２９６／２９７、８４／１４７／２９７／２９８／３０３、８４／１５６／１７３／２０４、８４／１７３／２０４／３０３、８４／１７３／２２４、８４／２２０／３１５、８４／２２４／２９７／２９８／２９９、８５、８６、８７、８７／１０６、８７／１０６／１１１／３１５、８７／１１１、８７／１１１／１５７／３１５、８７／１１１／２００、８７／１１１／２００／２４６、８７／１５７、８７／１５７／１８０／２００、８７／１５７／３１５、８７／３１５、８９、９１、９１／１０９／１１１、９１／１０９／１４７／２０５／２１０／２３４、９２、９２／１７３／２０４／２９０／３０３、９３、９４、９６、９７、９８、９８／１４３／１５８／１７０／１８５／２９６／２９７、９８／１４３／１６２／２６６、９８／１４３／１８５／２６６、９８／１６２／１９３、９８／１７０／１９３／１９７、９８／１８５、９８／１８５／１９３／１９７／２６６／２９７、９９、９９／１０３／２１９、９９／２３５、９９／２５６／３５１／３５５、１００、１００／１０１／２１１／２７８／３３８／３５０／３５３、１００／１１１、１００／１１１／３５３、１００／１２０／１９７／２４２、１００／２３５／２６８／３１５／３４６、１００／２３５／２６８／３４６／３５５、１００／２３５／２６８／３６６／３７３、１００／２３５／３４６／３５０／３５３、１００／２６８／３４６、１００／２６８／３６６、１０１、１０２、１０３、１０３／１０６／２０７／３２１、１０３／１０６／２１０、１０３／１０６／３６０、１０３／１１１／２３５／２８０／３６０、１０３／１１１／２３５／３２１／３２４、１０３／１６０／１８０、１０３／１６０／２９７、１０３／２０７／２１０／２３５／３２１、１０３／２１０／２３２／２３５、１０３／２１９／２５６、１０３／２１９／３３８／３５８、１０３／２３２／２７３、１０３／２３３、１０３／２５６、１０３／２５９、１０３／３２４、１０４、１０４／１０６、１０４／１０６／１１１／２００／２０１／２３５／２６８／３６８、１０４／１０６／１１１／２００／２０１／２６８、１０４／１０６／１１１／２０１／２３５／３６８、１０４／１０６／２００／２０１／２６８、１０４／１２０／２３２／３５３、１０４／１８２、１０４／２００／２０７／２３７／３４４／３７３／３８７、１０４／２０７／３４４／３８７、１０５、１０６、１０６／１１１、１０６／１１１／１５７、１０６／１１１／２００／２０１、１０６／１１１／２００／２０１／２６８／３６８、１０６／１１１／２００／２３５／３６８、１０６／１１１／２０１、１０６／１１１／２０１／３６８、１０６／１１１／３２２／３２４／３８６、１０６／１１１／３２２／３８３／３８６、１０６／１７３／２００／２３５／３１５、１０６／１７３／３８８、１０６／１８２／２０３／２２６／２３５／３４２／３４６、１０６／２００／２０１、１０６／２００／２０１／２０９／３６８、１０６／２００／２０１／２３５、１０６／２００／２０１／３６８、１０６／２００／２２６／３７３、１０６／２００／３２１／３２２／３８３、１０６／２０１／２３５、１０６／２０１／２６８、１０６／２０７／２１０／２３２／２３５／３２１／３６８／３７３、１０６／２０７／２３５／３２１／３６８、１０６／２２６／２３５／３７３、１０６／２３５、１０６／２３５／２７３／３７３、１０６／２３５／３８３／３８６、１０６／２７０／３４４、１０６／２７３、１０６／３１５、１０６／３２４、１０７、１０８、１０９、１０９／１１１／２０５／２１０／２３４／２８９、１１０、１１１、１１１／１５７／１８０／２００／３１５、１１１／１６０／２３３／２３５／２９７、１１１／１７３、１１１／１７３／２３５／３１５、１１１／１９７／２４２、１１１／２００／２０１／２６８、１１１／２００／３６８、１１１／２２６、１１１／２２６／３２１／３４４／３４６／３６９／３８７、１１１／２３５、１１１／２３５／２６８／３２７／３４６、１１１／２３５／２６８／３４６、１１１／２３５／２７１／３３９、１１１／２３５／２７２／３３９／３８６、１１１／２３５／２８０、１１１／２３５／３３９、１１１／２３５／３４６／３５０、１１１／２６８、１１１／２７２／３３９／３８６、１１１／３２２／３６０、１１１／３２７、１１１／３４６、１１２、１１３、１１３／３５５、１１５、１１６、１１７、１１８、１１９、１２０、１２２、１２３、１２４、１２５、１２８、１２８／１５９／２０９／３５２／３５３、１２８／２０９、１２８／２０９／２、１２８／２０９／２１９、１２８／２０９／２１９／３５２／３５３／３７１、１２８／２９６、１３１、１３３、１３３／１３５／３０７／３１５、１３３／２２０／２３３／２７１、１３３／３０７、１３４、１３４／３５３、１３５、１３６、１４０、１４０／１４１／２５６／２６１／３３９、１４０／１８０／２４４／２６１／３３９／３６０、１４０／１９２／１９３／３０２、１４０／１９３／３０２、１４１、１４１／２４４／２６１／３６０、１４１／２４４／３６０、１４１／２５６／３３９／３６０、１４１／２６１、１４１／２６１／３３９、１４１／３６０、１４３、１４３／１５８／１９７／２６６、１４３／２６６／２９６／２９７、１４４、１４４／２２０、１４５、１４７、１４７／１５８／１６２／１９０／２２４／２９６／２９７／２９９／３０３、１４７／１５８／１８２／２２０／２３３、１４７／１５８／１８２／２３０／２３３／２４９／２７１／３３９、１４７／１５８／１８２／２３３／２４９、１４７／１５８／１８２／２３３／２７１／３０７／３３９、１４７／１５８／１８２／２３３／３０７／３３９、１４７／１５８／１８２／２３３／３１５／３４５、１４７／１５８／１８２／２７１／３１５、１４７／１５８／１８２／３１５、１４７／１５８／２２０／２３０／２３３／２４９／３４５、１４７／１５８／２２０／２４９／３１５、１４７／１５８／２３３／２４９、１４７／１８２／２２０／２３３／３４５、１４７／１８２／２３０／３０７／３１５／３３９、１４７／１８２／２３３、１４７／１８２／２３３／２４９、１４７／１８２／２４９／２７１／３０７、１４７／１８２／３４５、１４７／２１０／２３４、１４７／２２０、１４７／２２４、１４７／２２４／２９７／２９８、１４７／２３２／２３３／２７１／３１５／３３９、１４７／２３３／３４５、１４７／２４９、１４７／２９０／３０３／３０７、１４７／３３９、１４８、１４９、１５０／２４４／２６１／３６０、１５１、１５３、１５４、１５４／１５６／２８２／３５０、１５４／１６６、１５４／１６６／２１０／２９６、１５４／１６６／３４６／３４７、１５４／１６６／３４７、１５４／１６７／２１０／３４７／３４９、１５４／１８４、１５４／１８７／２２０／３５０／３８６、１５４／２９３、１５４／２９６／３４７、１５４／３３９、１５４／３４７、１５４／３５０、１５４／３８６、１５５、１５５／２２０／３２５／３３９、１５６、１５６／１５８／１９０／１９３／３６４、１５６／１６６／１６７、１５６／２１０／３３９、１５７、１５７／１６０、１５７／１６２、１５７／１６２／２４１／２４２／２６０、１５７／１６２／２４１／２４２／３２７／３３１、１５７／１６２／２４１／２７８／３３１／３６８、１５７／１６２／２４１／３３１、１５７／１６２／２４２／２６０／３３１、１５７／１６２／３３１、１５７／３１５、１５７／３２７／３３１、１５７／３６８、１５８、１５８／１８２／２２０／２３０／２４９／３０７／３３９、１５８／１９３／２９０／３０３、１５８／２３３、１５８／３０７、１５８／３１５、１５８／３１５／３３９、１５９、１６０、１６０／２９６、１６１、１６１／１６２、１６１／２３７、１６２、１６２／１９３／２３２／２６７／３０２／３６０、１６２／２２４、１６２／２３７／２６５／２６６／２６７／３０２、１６２／２４１／２７８／３２７／３３１、１６２／２４１／３３１、１６２／２４２／３６８、１６２／２６５、１６２／２６７／３６０、１６２／３６７／３７１、１６３、１６３／２０１、１６４、１６４／２７５／２８０、１６５、１６６、１６６／２１０、１６６／２９６、１６６／３４６／３４７、１６７、１６７／２１０／３４６／３４９、１６８、１６９／３９０、１７０、１７０／１９３／１９７、１７３、１７３／２０４、１７３／２０４／３０３、１７３／２９７、１７４、１７５、１７６、１７７、１８０、１８０／３３９、１８１、１８２、１８２／１８６／２５６／３６０、１８２／１９７、１８２／２０１／２０３／２２６／２３４／２３５／３４６、１８２／２０１／２０３／２２６／２３４／３４２／３４６、１８２／２０１／２０３／２２６／２３５、１８２／２０１／２０３／２２６／２３５／３４６、１８２／２０１／２０３／２２６／３４２／３４６、１８２／２０１／２０３／２６８／３４６、１８２／２０１／２２６／２３４／３４２／３４６、１８２／２０１／２２６／２３５／３４６、１８２／２０１／ ２２６／２６８／３４６、１８２／２０１／３４２、１８２／２０３／２２６、１８２／２０７／２７９／３６８、１８２／２２０／３４５、１８２／２２６／２３４／２６８／３４２／３４６、１８２／２２６／２３４／３４６、１８２／２２６／２３５、１８２／２２６／２３５／２６８／３４６、１８２／２２６／２３５／３４２、１８２／２２６／２３５／３４６、１８２／２２６／２６８／３４２／３４６、１８２／２２６／３４２、１８２／２２６／３４６、１８２／２３０、１８２／２３０／２３３、１８２／２３４／３８０、１８２／２５６、１８２／２７９／３６８／３８０、１８２／２９８／３６４、１８２／３４２／３４６、１８２／３４５、１８４、１８４／１８７、１８４／１８８／２００／２０３／２１１／２４２、１８４／１８８／２００／２０３／２４２／２９７、１８４／１８８／２００／２０３／２９０／２９７／３６８、１８４／１８８／２０３／２９０／２９７、１８４／１８８／２１１／２４２／２９０／３６８、１８４／１８９、１８４／１８９／２０６／２９７／３６８、１８４／１８９／２９７、１８４／２００、１８４／２０６／２４２／２９０／２９７、１８４／２１１、１８４／２４２、１８４／２４２／２９７／３６８、１８４／２９０／２９７、１８４／２９０／３６８、１８４／２９３、１８５、１８６、１８６／２５６、１８６／２５６／２７０、１８７、１８７／２２０／３５０、１８７／２９３、１８８、１８８／２１１、１８８／２１１／２４２／２９０／２９７／３６８、１８９、１８９／２００、１９０、１９０／１９３／１９７、１９１、１９２、１９２／１９３、１９２／１９３／２１１／２４２／２９７／３６８、１９２／１９３／２９０／２９７／３６８、１９３、１９３／１９４、１９３／１９７／２９６／３０３、１９３／２６７、１９３／２９０、１９４、１９４／２４２、１９４／２４２／２９０、１９５、１９６、１９７、１９７／２９８、１９８、１９８／２８９、１９９、２００、２００／２０２／２０３、２００／２０３、２００／２０３／２２６／２３０／３３９、２００／２０３／２２９／２３４、２００／２０３／２９０、２００／２０３／２９０／３６８、２００／２０６、２００／２２６、２００／２２６／３２１／３７３、２００／２２６／３７３、２００／２３０／２５７、２００／２３４／２９０、２００／２３５／３６８、２００／２７０、２００／２７０／２７５／３３９、２００／２７０／３２２／３８３、２００／２７１、２００／２９０／２９２／３６０、２００／３４４、２００／３６８、２０１、２０１／２０２、２０１／２０２／２７２／３６０、２０１／２０３、２０１／２０３／２２６／２３４、２０１／２０３／２２６／２３４／３４２／３４６、２０１／２０３／２２６／２３４／３４６、２０１／２０３／２２６／２６８／３４２、２０１／２０３／２２６／３４６、２０１／２０３／２６８／３４２、２０１／２０３／２６８／３４６、２０１／２２６／２３４／３４２／３４６、２０１／２２６／２３４／３４６、２０１／２２６／２３５／３４２、２０１／２２６／２６８／３４６、２０１／２２６／３４６、２０１／２３０／２５７、２０１／２３０／２９０、２０１／２３５／２６８／３４２／３４６、２０１／３６８、２０１／３７１、２０２、２０２／２３３、２０２／２３５／３６０／３６７／３７１、２０３、２０３／２０６／３６８、２０３／２２６、２０３／２２６／２３０、２０３／２２６／２３０／２５７、２０３／２２６／２３４／２３５、２０３／２２６／２３４／２３５／３４６、２０３／２２６／２３５／２６８／３４６、２０３／２２６／２３５／３４２、２０３／２２６／２３５／３４６、２０３／２２６／３４２／３４６、２０３／２２６／３４６、２０３／２２９、２０３／２４２／２９０／２９７／３６８、２０３／２９７、２０４、２０４／２６４／３４０、２０５、２０６、２０６／２９０、２０６／２９７、２０７、２０７／２１０／２７３／２７９／３４４、２０７／２１０／２７３／２７９／３４４／３４９／３７３、２０７／２１０／２７３／３２１／３４４／３４９、２０７／２３３／２３７／３４４／３８７、２０７／２３５、２０７／２３５／３２７／３６０／３７１、２０７／２３５／３６８、２０７／２７１／３６８／３８０、２０７／２７３、２０７／２７９／３４９、２０７／３６８／３７３、２０７／３７３、２０７／３８０、２０７／３８７、２０９、２０９／２１１、２１０、２１０／２１１、２１０／２１１／２４２、２１０／２７３／２７９、２１０／２７３／２７９／３４９、２１０／２７９、２１０／３３９、２１１、２１１／３６８、２１７、２１９、２１９／３００、２１９／３５８、２２０、２２０／２２４／３１５、２２０／２４９、２２０／２９３、２２０／３０７／３３９、２２０／３１５、２２０／３３９、２２０／３４９、２２４、２２４／２９６／２９８、２２４／２９７／２９９、２２４／２９８／２９９、２２６、２２６／２３４／２３５／３４６、２２６／２３４／３４２、２２６／２３５／２６８、２２６／２３５／２６８／３４２／３４６、２２６／２３５／２６８／３４６、２２６／２３５／２７２／３７３、２２６／２３５／３４２、２２６／２３５／３４６、２２６／２６８、２２６／２６８／３４２、２２６／２６８／３４２／３４６、２２６／２６８／３４６、２２６／２７２／３７３、２２６／３２１／３６９、２２６／３４２、２２６／３４２／３４６、２２６／３４６、２２６／３６６、２２６／３６６／３６９／３８７、２２７、２２８、２２９、２２９／３２５、２２９／３６０、２３０、２３１、２３２、２３２／２３５、２３２／２３５／２７３／３６８、２３２／２３５／３２１、２３２／３４６／３５０／３５５、２３３、２３３／２７１、２３３／３１５、２３３／３６７／３７１、２３３／３７１、２３４、２３５、２３５／２６８／３２７／３４６、２３５／２６８／３４６、２３５／２７１／２７２／３３９、２３５／２７１／２７２／３３９／３８６、２３５／２７１／３３９、２３５／２７１／３３９／３８６、２３５／２７２、２３５／２７２／３３９、２３５／２７２／３８６、２３５／２７３、２３５／２８０、２３５／２８０／３２１／３２２／３２４、２３５／２８０／３２１／３２４／３８３／３８６、２３５／３１５、２３５／３１５／３５３／３５５、２３５／３３９、２３５／３３９／３８６、２３５／３４６、２３５／３４６／３５０、２３５／３５３、２３５／３５３／３５５、２３６、２３７、２３７／２６５、２３７／２６６／３６０、２３７／２７１、２３７／３６０、２３７／３８１、２３８、２３９、２４１、２４１／２４２／３６８、２４２、２４２／２４４、２４２／２９０、２４２／２９０／２９７、２４２／３３１、２４３、２４４、２４４／２５６／２６１／３３９／３６０、２４５、２４６、２４７、２４８、２４９、２４９／３１５／３３９、２５０、２５２、２５３、２５６、２５６／２５９、２５６／３６０、２５７、２５８、２５９、２５９／２７６／３８７、２６０、２６０／３２７、260 / 331, 260 / 368, 261, 262, 262 / 325 / 349, 263, 264, 265, 265 / 346, 266, 266 / 297 / 299, 267, 267 / 270 / 275 / 339 / 347, 267 / 272 / 275, 268, 268 / 315 / 327 / 346, 268 / 315 / 346, 268 / 315 / 346 / 366, 268 / 315 / 355, 268 / 324, 268 / 324 / 327 / 346, 268 / 327 / 346, 268 / 346, 268 / 346 / 350 / 353 / 355, 268 / 353, 268 / 355 / 366, 269, 270, 270 / 309, 270 / 322 / 344 / 383, 271, 271 / 339, 271 / 380, 272, 273, 273 / 279, 273 / 279 / 373, 274, 275, 275 / 291, 276, 277, 278, 279, 280, 280 / 383, 282, 284, 286, 287, 288, 289, 290, 290 / 303 / 364, 291, 292, 293, 293 / 350, 293 / 350 / 386, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 313, 314, 315, 315 / 324 / 327 / 355 / 366, 315 / 327 / 346, 319, 321, 321 / 324, 322, 324, 324 / 327 / 346, 324 / 346 / 350, 324 / 346 / 355 / 366, 324 / 355, 325, 325 / 339, 326, 327, 327 / 346, 328, 330, 331, 332, 338, 339, 341, 342, 342 / 363, 344, 344 / 370, 345, 346, 347, 348, and / or any combination thereof, wherein:

2. The engineered terminal deoxynucleotidyl transferase of claim 1, wherein the amino acid positions of the polypeptide sequence are numbered with reference to SEQ ID NOs: 5296, 5628, 5630, 5632, and / or 5636.

17. 10. The engineered terminal deoxynucleotidyl transferase of claim 1, having at least one improved property compared to a wild-type terminal deoxynucleotidyl transferase or a reference engineered terminal deoxynucleotidyl transferase.

18. 18. The engineered terminal deoxynucleotidyl transferase of claim 17, having at least one improved property compared to a wild-type terminal deoxynucleotidyl transferase or a reference engineered terminal deoxynucleotidyl transferase, wherein the improved property is selected from increased thermostability, increased activity at elevated temperatures, increased soluble expression, decreased by-product formation, and increased conversion of substrate to product.

19. The engineered terminal deoxynucleotidyl transferase of claim 18, wherein the engineered terminal deoxynucleotidyl transferase comprises increased soluble expression compared to a wild-type terminal deoxynucleotidyl transferase or a reference engineered terminal deoxynucleotidyl transferase.

20. The engineered terminal deoxynucleotidyl transferase of claim 18, wherein the engineered terminal deoxynucleotidyl transferase comprises increased activity at elevated temperatures compared to a wild-type terminal deoxynucleotidyl transferase or a reference engineered terminal deoxynucleotidyl transferase.

21. The engineered terminal deoxynucleotidyl transferase of claim 18, wherein the engineered terminal deoxynucleotidyl transferase comprises reduced by-product formation compared to a wild-type terminal deoxynucleotidyl transferase or a reference engineered terminal deoxynucleotidyl transferase.

22. The engineered terminal deoxynucleotidyl transferase of claim 18, wherein the engineered terminal deoxynucleotidyl transferase comprises increased conversion of substrate to product compared to a wild-type terminal deoxynucleotidyl transferase or a reference engineered terminal deoxynucleotidyl transferase.

23. The engineered terminal deoxynucleotidyl transferase of claim 18, wherein the engineered terminal deoxynucleotidyl transferase comprises increased thermal stability compared to a wild-type terminal deoxynucleotidyl transferase or a reference engineered terminal deoxynucleotidyl transferase.

24. 24. The engineered terminal deoxynucleotidyl transferase of claim 23, wherein the improved property is increased thermal stability of 2-fold, 5-fold, 10-fold, 15-fold, or more, compared to a reference terminal deoxynucleotidyl transferase.

25. 21. The engineered terminal deoxynucleotidyl transferase of claim 20, wherein the improved property is an increased activity of the engineered TdT at 40°C, 45°C, 50°C, 55°C, 60°C, 65°C, 70°C, or 75°C that is 2-fold, 5-fold, 10-fold, 15-fold, or more, compared to a reference terminal deoxynucleotidyl transferase.

26. 26. The engineered terminal deoxynucleotidyl transferase of any of claims 19 and 23-25, wherein the engineered terminal deoxynucleotidyl transferase comprises at least one residue difference at an amino acid position selected from 80, 87, 203, 111, 143, 147, 157, 160, 180, 209, 226, 256, 261, 266, 279, 327, 339, 349, 353, and 364.

27. 27. The engineered terminal deoxynucleotidyl transferase of claim 26, wherein the engineered terminal deoxynucleotidyl transferase comprises at least one residue difference selected from 143A, 157V, 203D, 209E, and 353N.

28. 19. The engineered terminal deoxynucleotidyl transferase of claim 18, wherein the improved property is increased conversion of the substrate to product by the engineered TdT to a rate of 5%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95% or higher.

29. 30. The engineered terminal deoxynucleotidyl transferase of any of claims 21, 22, and 28, wherein the engineered terminal deoxynucleotidyl transferase comprises at least one residue difference at an amino acid position selected from 53, 65, 159, 211, 217, 224, 271, 272, 273, 275, 278, 331, 341, and 391.

30. 30. The engineered terminal deoxynucleotidyl transferase of claim 29, wherein the engineered terminal deoxynucleotidyl transferase comprises at least one residue difference selected from 53T, 159R, 211V, 217R, 271H, 272A, 275Q, 278G, 331K, and 391L.

31. 2. The engineered terminal deoxynucleotidyl transferase of claim 1, wherein the at least one residue difference or set of residue differences comprises a residue difference or set of residue differences at an amino acid position selected from 53, 65, 80, 87, 111, 143, 147, 157, 159, 160, 180, 203, 209, 211, 217, 224, 226, 256, 261, 266, 271, 272, 273, 275, 278, 279, 327, 331, 339, 341, 349, 364, and 391, and / or any combination thereof, wherein the amino acid positions are numbered with reference to SEQ ID NOs: 5296, 5628, 5630, 5632, and / or 5636.

32. 10. The engineered terminal deoxynucleotidyl transferase of claim 1, which is purified.

33. 2. A polynucleotide sequence encoding at least one engineered terminal deoxynucleotidyl transferase according to claim 1.

34. 1. A polynucleotide sequence comprising at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or greater sequence identity to a reference sequence of SEQ ID NOs: 5295, 5627, 5629, 5631, and / or 5635 and / or a functional fragment thereof, wherein said polynucleotide sequence encodes an engineered polypeptide comprising at least one residue difference at one or more amino acid positions.

35. 34. The polynucleotide sequence of claim 33, wherein the polynucleotide sequence encodes at least one engineered terminal deoxynucleotidyl transferase comprising a sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or greater sequence identity to a reference sequence of SEQ ID NO: 5296, 5628, 5630, 5632, and / or 5636.

36. 36. The polynucleotide sequence of claim 35, wherein the sequence comprises SEQ ID NO: 5295, 5627, 5629, 5631, and / or 5635.

37. The polynucleotide sequence of claim 33, wherein the polynucleotide sequence is operably linked to a control sequence.

38. The polynucleotide sequence of claim 37, wherein the polynucleotide sequence is codon-optimized.

39. 34. An expression vector comprising at least one polynucleotide sequence according to claim 33.

40. 40. A host cell transformed with at least one expression vector according to claim 39.

41. 10. A composition comprising at least one engineered terminal deoxynucleotidyl transferase according to claim 1.

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