Orthogonal aminoacyl TRNA synthetase pairs for protein production

Abstract

The present disclosure relates to orthogonal transfer RNA and aminoacyl tRNA synthetase pairs that can be used in a heterologous host cell to produce a protein of interest.

Description

[0001] ORTHOGONAL AMINOACYL TRNA SYNTHETASE PAIRS FOR PROTEIN

[0002] PRODUCTION

[0003] RELATED APPLICATION

[0004] This application claims the benefit under 35 U.S.C. § 119(e) of U.S. provisional application number 63 / 575,451, filed April 5, 2024, which is incorporated by reference herein in its entirety.

[0005] REFERENCE TO AN ELECTRONIC SEQUENCE LISTING

[0006] The content of the electronic sequence listing (H049870821WO00-SEQ-HJD.xml; Size: 1,854,320 bytes; and Date of Creation: March 31, 2025) is herein incorporated by reference in its entirety.

[0007] BACKGROUND

[0008] The human population is estimated to reach 9.7 billion by 2050, contributing to an increased global demand for new medicines and antibiotics, housing, and human-made materials, such as plastic, asphalt, aluminum, and cement. There is a pressing need to develop greener alternatives to materials production and replace materials produced from fossil-fuels with more sustainable alternatives. In addition, development of next- generation smallmolecule synthesis approaches enabled by new biocatalysts could accelerate the search for cures and treatments of disease.

[0009] Synthetic chemical approaches to polymer production provide access to a broad range of chemical structures yet are not environmentally sustainable as they rely on petroleum production. In addition, synthetic materials are constrained by the lack of sequence-defined polymerization methods. In contrast, living systems employ sequence-defined processes to synthesize biomolecules in which the molecular information encoded by nucleic acids is converted into sequence-defined protein polymers. Evolution has designed natural biopolymers with a range of strength, elasticity, and stability, such as elastin, collagen, silk and keratin optimized for specialized function in nature.

[0010] SUMMARY

[0011] Aspects of the present disclosure relate to orthogonal transfer RNA (tRNA) and aminoacyl tRNA synthetase (aaRS) (aaRS-tRNA) pairs that can be used in a heterologous host cell (e.g., heterologous to the cell from which the aaRS-tRNA pairs are obtain) to produce a protein of interest.

[0012] Aspects of the present disclosure relate to a host cell comprising: a transfer ribonucleic acid (tRNA); and an aminoacyl-tRNA synthetase (aaRS); wherein the tRNA and aaRS are from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilius, H. mukohataei DSM 12286, or M. bourgensis, and the tRNA and aaRS are heterologous to the host cell.

[0013] In some embodiments, the tRNA comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 1-727. In some embodiments, the aaRS comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 728-1454. In some embodiments, the tRNA comprises an amino acid sequence having at least 90% or at least 95% identity to any one of SEQ ID NOs: 1-727 and the aaRS comprises an amino acid sequence having at least 90% or at least 95% identity to any one of SEQ ID NOs: 728-1454.

[0014] Aspects of the present disclosure relate to a host cell comprising: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1- 727; and an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728-1454; wherein the tRNA and aaRS are heterologous to host cell.

[0015] Aspects of the present disclosure relate to a host cell comprising: a transfer ribonucleic acid (tRNA) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454, optionally wherein the tRNA and aaRS are heterologous to host cell.

[0016] In some embodiments, the tRNA is charged with a standard amino acid. In some embodiments, the tRNA is charged with a nonstandard amino acid. In some embodiments, the nonstandard amino acid is cystine, desmosine, isodesmosine, hydroxyproline, hydroxylysine, gamma-carboxyglutamate, phosphoserine, phosphothreonine, phosphotyrosine, N-acetyl lysine, methyllysine, para-azido-phenylalanine, propargyloxy-phenylalanine, benzoylphenylalanine, or dibenzocyclooctane-lysine.

[0017] In some embodiments, the host cell is a prokaryotic cell. In some embodiments, the prokaryotic cell is a bacterial cell or an archaebacterial cell. In some embodiments, the bacterial cell is an Escherichia coli (E. coli) cell. In some embodiments, the host cell is a eukaryotic cell. In some embodiments, the eukaryotic cell is an animal cell, an insect cell, a fungal cell, or an algal cell. In some embodiments, the animal cell is a mammalian cell. In some embodiments, the mammalian cell is a human cell. In some embodiments, the insect cell is an Sf9 cell, a Tni cell, an Sf21 cell, or an S2 cell.

[0018] Aspects of the present disclosure relate to a cell-free system, comprising: a transfer ribonucleic acid (tRNA); and an aminoacyl-tRNA synthetase (aaRS); wherein the tRNA and aaRS are from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilius, H. mukohataei DSM 12286, or M. bourgensis, and the tRNA and aaRS are heterologous to the cell-free system.

[0019] In some embodiments, the tRNA comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 1-727. In some embodiments, the aaRS comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 728-1454. In some embodiments, the tRNA comprises an amino acid sequence having at least 90% or at least 95% identity to an amino acid sequence of any one of SEQ ID NOs: 1-727 and an aaRS comprising an amino acid sequence having at least 90% or at least 95% identity to any one of SEQ ID NOs: 728-1454.

[0020] Aspects of the present disclosure relate to a cell-free system comprising: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1- 727; and an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728-1454; wherein the tRNA and aaRS are heterologous to cell-free system.

[0021] Aspects of the present disclosure relate to a cell-free system comprising: a transfer ribonucleic acid (tRNA) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454, optionally wherein the tRNA and aaRS are heterologous to cell-free system.

[0022] In some embodiments, the tRNA is charged with a standard amino acid. In some embodiments, the tRNA is charged with a nonstandard amino acid. In some embodiments, the nonstandard amino acid is cystine, desmosine, isodesmosine, hydroxyproline, hydroxylysine, gamma-carboxyglutamate, phosphoserine, phosphothreonine, phosphotyrosine, N-acetyl lysine, methyllysine, para-azido-phenylalanine, propargyloxy-phenylalanine, benzoylphenylalanine, or dibenzocyclooctane-lysine.

[0023] In some embodiments, the cell-free system is a lysate. In some embodiments, the lysate is derived from an organism. In some embodiments, the organism is a plant, a human, a fungus, an insect, or a bacterium. In some embodiments, the bacterium is cyanobacteria or Vibrio natriegens.

[0024] Aspects of the present disclosure relate to a composition comprising: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1- 727; an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728-1454; and a nonstandard amino acid.

[0025] Aspects of the present disclosure relate to a composition comprising: a transfer ribonucleic acid (tRNA) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1-727; an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454; and optionally a nonstandard amino acid.

[0026] In some embodiments, the tRNA is charged with a standard amino acid. In some embodiments, the tRNA is charged with a nonstandard amino acid. In some embodiments, the nonstandard amino acid is cystine, desmosine, isodesmosine, hydroxyproline, hydroxylysine, gamma-carboxyglutamate, phosphoserine, phosphothreonine, phosphotyrosine, N-acetyl lysine, methyllysine, para-azido-phenylalanine, propargyloxy-phenylalanine, benzoylphenylalanine, or dibenzocyclooctane-lysine.

[0027] Aspects of the present disclosure relate to a method of producing a protein of interest, comprising: delivering to a host cell a transfer ribonucleic acid (tRNA) and an aminoacyl- tRNA synthetase (aaRS), wherein the tRNA and aaRS are from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilius, H. mukohataei DSM 12286, or M. bourgensis, and the tRNA and aaRS are heterologous to the host cell; and optionally delivering to the host cell a nucleic acid encoding a protein of interest.

[0028] In some embodiments, the tRNA comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 1-727. In some embodiments, the aaRS comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 728-1454. In some embodiments, the tRNA comprises an amino acid sequence having at least 90% or at least 95% identity to an amino acid sequence of any one of SEQ ID NOs: 1-727 and an aaRS comprising an amino acid sequence having at least 90% or at least 95% identity to any one of SEQ ID NOs: 728-1454.

[0029] Aspects of the present disclosure relate to a method of producing a protein of interest, comprising: delivering to a host cell: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728-1454, wherein the tRNA and aaRS are heterologous to the host cell; and optionally delivering to the host cell a nucleic acid encoding a protein of interest.

[0030] Aspects of the present disclosure relate to a method of producing a protein of interest, comprising: delivering to a host cell: a transfer ribonucleic acid (tRNA) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454, optionally wherein the tRNA and aaRS are heterologous to the host cell; and optionally delivering to the host cell a nucleic acid encoding a protein of interest.

[0031] In some embodiments, the tRNA is charged with a standard amino acid. In some embodiments, the tRNA is charged with a nonstandard amino acid. In some embodiments, the nonstandard amino acid is cystine, desmosine, isodesmosine, hydroxyproline, hydroxylysine, gamma-carboxyglutamate, phosphoserine, phosphothreonine, phosphotyrosine, N-acetyl lysine, methyllysine, para-azido-phenylalanine, propargyloxy-phenylalanine, benzoylphenylalanine, or dibenzocyclooctane-lysine.

[0032] In some embodiments, the host cell is a prokaryotic cell. In some embodiments, the prokaryotic cell is a bacterial cell or an archaebacterial cell. In some embodiments, the bacterial cell is an Escherichia coli (E. coli) cell. In some embodiments, the host cell is a eukaryotic cell. In some embodiments, the eukaryotic cell is selected from animal cells, insect cells, fungal cells, and algal cells. In some embodiments, the animal cells are mammalian cells. In some embodiments, the mammalian cells are human cells.

[0033] Aspects of the present disclosure relate to a method of producing a protein of interest, comprising: delivering to a lysate a transfer ribonucleic acid (tRNA) and an aminoacyl-tRNA synthetase (aaRS), wherein the tRNA and aaRS are from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilius, H. mukohataei DSM 12286, or M. bourgensis, and the tRNA and aaRS are heterologous to the lysate; and optionally delivering to the lysate a nucleic acid encoding a protein of interest.

[0034] In some embodiments, the tRNA comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 1-727. In some embodiments, the aaRS comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 728-1454. In some embodiments, the tRNA comprises an amino acid sequence having at least 90% or at least 95% identity to an amino acid sequence of any one of SEQ ID NOs: 1-727 and an aaRS comprising an amino acid sequence having at least 90% or at least 95% identity to any one of SEQ ID NOs: 728-1454.

[0035] Aspects of the present disclosure relate to a method of producing a protein of interest, comprising: delivering to a lysate: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728-1454, wherein the tRNA and aaRS are heterologous to the lysate; and optionally delivering to the lysate a nucleic acid encoding a protein of interest.

[0036] Aspects of the present disclosure relate to a method of producing a protein of interest, comprising: delivering to a lysate: transfer ribonucleic acid (tRNA) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1- 727; and an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454, optionally wherein the tRNA and aaRS are heterologous to the lysate; and optionally delivering to the lysate a nucleic acid encoding a protein of interest.

[0037] In some embodiments, the tRNA is charged with a standard amino acid. In some embodiments, the tRNA is charged with a nonstandard amino acid. In some embodiments, the nonstandard amino acid is cystine, desmosine, isodesmosine, hydroxyproline, hydroxylysine, gamma-carboxyglutamate, phosphoserine, phosphothreonine, phosphotyrosine, N-acetyl lysine, methyllysine, para-azido-phenylalanine, propargyloxy-phenylalanine, benzoylphenylalanine, or dibenzocyclooctane-lysine.

[0038] In some embodiments, the lysate is derived from an organism. In some embodiments, the organism is a plant, a human, a fungus, an insect, or a bacterium. In some embodiments, the bacterium is cyanobacteria or Vibrio natriegens.

[0039] Aspects of the present disclosure relate to a method of evolving a tRNA-aaRS pair, comprising: evolving a native transfer ribonucleic acid (tRNA) to accept a nonstandard amino acid; and evolving a native aminoacyl-tRNA synthetase (aaRS) to charge the tRNA with the nonstandard amino acid; the evolved tRNA comprises an amino acid sequence having greater than 75% and less than 100% identity to any one of SEQ ID NOs: 1-727, and wherein the evolved aaRS comprises an amino acid sequence having greater than 75% and less than 100% identity to any one of SEQ ID NOs: 728-1454.

[0040] In some embodiments, the native tRNA comprises an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 1-727, and wherein the native aaRS comprises an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 728-1454.

[0041] Aspects of the present disclosure relate to a method comprising: combining a cell lysate that comprises one or more native aminoacyl tRNA synthetases (aaRSs) with a pool of in vitro transcribed transfer ribonucleic acid (tRNA) molecules; extracting one or more of the synthetic tRNA molecules from the cell lysate to obtain isolated tRNA molecules; and assessing aminoacylation of one or more of the isolated tRNA molecules using periodate oxidation and beta-elimination.

[0042] In some embodiments, the method further comprises linking adaptors to one or more of the isolated tRNA molecules to produce adaptor- linked tRNA molecules. In some embodiments, the method further comprises reverse transcribing one or more of the adaptor- linked tRNA molecules to produce one or more DNA molecules. In some embodiments, the method further comprises sequencing one or more of the DNA molecules, optionally using a next generation sequencing (NGS) technology.

[0043] In some embodiments, the method further comprises assessing aminoacylation of one or more of the isolated tRNA molecules, optionally wherein the assessing comprises determining whether the one or more of the isolated tRNA molecules comprises a 3 '-terminal CCA sequence or is missing a 3 '-terminal adenosine. In some embodiments, the method further comprises quantifying a ratio of the isolated tRNA molecules comprising a 3 '-terminal CCA sequence to the isolated tRNA molecules missing a 3 '-terminal adenosine, thereby quantifying a percentage of aminoacylation for the isolated tRNA molecules.

[0044] Aspects of the present disclosure relate to a method, comprising: combining: a cell lysate that comprises one or more native aminoacyl tRNA synthetases (aaRSs); a doublestranded DNA encoding a heterologous aaRS; a heterologous transfer ribonucleic acid (tRNA); and a detectably labeled adenosine triphosphate (ATP); and assessing aminoacylation of the tRNA.

[0045] In some embodiments, the tRNA is produced in the cell by in vitro transcription. In some embodiments, the detectably labeled ATP is isotopically labeled. In some embodiments, the assessing is executed using liquid chromatography-mass spectrometry (LC-MS). In some embodiments, the method is automated. In some embodiments, the cell lysate is obtained from prokaryotic cells, optionally Escherichia coli cells. In some embodiments, the cell lysate is obtained from eukaryotic cells. In some embodiments, the eukaryotic cells are selected from insect cells, plant cells, and mammalian cells, optionally human cells. BRIEF DESCRIPTION OF THE DRAWINGS

[0046] The accompanying drawings are not intended to be drawn to scale. In the drawings, each identical or nearly identical component that is illustrated in various figures is represented by a like numeral. For purposes of clarity, not every component may be labeled in every drawing. In the drawings:

[0047] FIG. 1 is a schematic of an exemplary method described herein.

[0048] FIG. 2 shows an overview of a transfer RNA (tRNA)-aminoacyl tRNA synthetase (aaRS) (aaRS-tRNA) translation system evolution approach.

[0049] FIG. 3 shows a workflow for high-throughput aminoacylation profiling.

[0050] FIG. 4 shows a workflow for use of Next-generation sequencing (NGS) to identify charged tRNAs.

[0051] FIG. 5 shows determination of tRNA charging in M. jannaschii tRNAs.

[0052] FIG. 6 shows determination of tRNA charging in M. jannaschii tRNAs.

[0053] FIG. 7 shows results of in vitro transcription and translation from a high throughput aminoacylation assay.

[0054] FIG. 8 shows assaying aminoacylation of E. coli tRNAtyrwith tyrosine by in vitro- produced tyrRS using LC-MS.

[0055] FIG. 9 shows determination by LC-MS of orthogonality between known orthogonal aaRStRNA pairs from M. bakeri and M. mazei, as well as the ability to charge multiple nsAAs.

[0056] FIG. 10 shows an aminoacylation assay set up robotically on E. coli tRNAtyr with tyrosine by in vztro-produced tyrRS using LC-MS.

[0057] FIG. 11 shows a workflow for detection of orthogonal tRNA aminoacylation with native tRNA modifications in lysates using isotopically labeled ATP and LC-MS.

[0058] FIG. 12 shows amino-acid adenylates from both orthogonal tRNAs and native tRNAs share the same retention times by LC-MS.

[0059] FIGs. 13A-13B shows discovery of orthogonal aaRS-tRNA pairs. Reactions were performed with (FIG. 13A) or without (FIG. 13B) double-stranded DNA (dsDNA) for the aaRS present in the reaction.

[0060] FIG. 14 shows an overview of expanding the number of aaRSs that can charge CGA- tRNAs.

[0061] DETAILED DESCRIPTION The development of biomaterials that can replace existing petroleum-based polymers holds promise for making a more sustainable future for humanity. In addition, the generation of novel sequence-defined materials holds promise for discovering novel classes of materials with improved mechanical, chemical, or biological properties. While natural enzymes composed of the twenty canonical L-a amino acids catalyze multiple classes of reactions, enzymes function predominantly in aqueous solvents, and their chemical repertoire is substantially narrower than of synthetic chemistry. By producing polypeptides containing nonstandard amino acid (nsAA) sidechains in precise positions, or biopolymers with nonpeptide backbones, it is possible to generate sequence-defined polymers that may function as catalysts, which can be evolved to achieve desired functions, and may exceed conventional chemical transformations in yield, purity, stereospecificity.

[0062] The field of genetic code expansion has opened the possibility to overcome the sparse chemical space afforded by natural translation and to allow control over sequences, unlike with synthetic materials. Previous work has shown site-specific incorporation of more than 150 nsAAs into proteins using orthogonal translations systems (OTSs). Photocaged, fluorescent, and bioorthogonal reactive monomers have provided new ways to study protein structure, dynamics, and post-translational modifications. Further, the generation of proteins with nonnatural chemistry has enabled the synthesis of novel therapeutics and genetically encoded materials.

[0063] While more than 150 nsAAs have been incorporated into proteins using genetic code expansion, the ability to produce multi-site incorporation of more than one nsAA into proteins or peptides in high purity is a major limitation for the field. To develop proteins with multi-site incorporation of nsAAs, multiple mutually orthogonal transfer RNA (tRNA)- aminoacyl transfer RNA synthetase (aaRS) pairs are necessary. Living cells have been repurposed for nsAA incorporation via open codons through whole genome recoding. This allowed the demonstration of multi-site incorporation of nsAAs. To introduce a greater number of nsAAs to the alphabet of translation, more codons will need to be recoded. Genome editing is a laborious and complex task, however, requiring multiple years of effort. In parallel work, flexizymes have been developed for charging nsAAs to tRNAs in vitro. Combined with the PURE translation system with reconstituted components, this allows translation of proteins with multiple nsAAs by removing some tRNAs and substituting them with flexizyme-charged tRNAs sharing the same anticodons. Nonetheless, aaRS charging of tRNAs even in lysates could be more scalable, as aaRSs can continuously charge tRNAs and translation reactions can be performed in cellular lysates, not only a reconstituted translation system with purified components. The combination of these factors can allow for larger throughput in production of proteins functionalized with nsAAs, allowing for production of new therapeutics and protein biomaterials.

[0064] Accordingly, aspects of the present disclosure relate to mutually orthogonal aaRSs and tRNAs that were generated for complete repurposing of all sense codons within E. coli or other translation systems. A method was developed to screen tRNA charging status and then to produce aaRSs in vitro and perform automated robotic in vitro aminoacylation reactions. Using these new methods, -10,000 tRNA sequences were assessed and new orthogonal tRNA-aaRS pairs discovered, which were validated to be functional by high resolution LC- MS. The tRNA-aaRS pairs discovered as discussed herein can be used, for example, to translate proteins and peptides with multi-site incorporation of nsAAs, with applications to new catalysis, biomaterials, and therapeutics.

[0065] Transfers RNAs and Aminoacyl tRNA Synthetases

[0066] Transfer ribonucleic acid (tRNA) is an adaptor molecule composed of ribonucleic acid (RNA). In eukaryotes, tRNA is typically between 76 and 90 nucleotides in length and it serves as the physical link between messenger RNA (mRNA) and the amino acid sequence of proteins. tRNA serves as a physical link between mRNA and amino acid sequences of proteins by carrying an amino acid to the protein- synthesizing machinery of a cell (ribosomes). tRNAs have a 3-nucleotide anticodon that is complementary to a 3-nucleotide codon in mRNA. mRNA comprising several 3-nucleotide codons binds to a ribosome. tRNAs carrying various amino acids then identify mRNA codons using their 3-nucleotide anticodon. Upon identification, the amino acid of the tRNA is removed from the tRNA and added to a growing chain of amino acids. Accordingly, tRNAs are essential to the translation or mRNA into protein. tRNA molecules have a primary structure comprised of nucleotides, a secondary structure in which the nucleotides form a cloverleaf structure, and a tertiary structure in which the secondary structure forms an L-shaped 3D structure that allows tRNA molecules to fit into specific sites of the ribosome.

[0067] Generally, tRNA molecules are defined by six regions: the acceptor stem, the CCA tail, the D loop, the anticodon loop, the T C loop, and the variable loop. The acceptor stem is a 7-9 base pair stem made by the base pairing of the 5 ’-terminal nucleotide of the tRNA with the 3’-nterminal nucleotide. The CCA tail is a cytosine-cytosine-adenine sequence at the 3’ end of the tRNA molecule. The D loop is a 4-6 base pair stem ending in a loop that often contains dihydrofluoridine. The anticodon loop is a 5 base pair stem that contains the 3- nucleotide anticodon. The T C loop is a location on the tRNA molecule that contains a modified uridine. The variable loop is a region on the tRNA that is variable in length. tRNA molecules with a variable loop with 10 or fewer base pairs are referred to as Class I tRNA molecules. tRNA molecules with a variable loop with more than 10 base pairs are referred to as Class II tRNA molecules. tRNA anticodons correspond to specific mRNA codons such that an mRNA sequence is translated into a pre-determined polypeptide chain. tRNA molecules, however, can be modified such that a tRNA with an anticodon for a stop codon instead delivers an amino acid. Modifying tRNA molecules in this way can allow for the production of unique polypeptide chains. tRNA molecules can be further modified such that the tRNA molecules deliver nonstandard amino acids (z.e., amino acids other than the 20 essential amino acids). tRNA molecules are charged with an amino acid such that the tRNA delivers a specific amino acid to a ribosome, the tRNA anticodon identifies an mRNA codon, then ribosome machinery removes the amino acid from the tRNA and adds the amino acid to a growing polypeptide chain. tRNA molecules are charged by aminoacyl tRNA synthetase (aaRS). aaRS is an enzyme that attaches the appropriate amino acid onto its corresponding tRNA. Each aaRS corresponds to a tRNA such that each amino acid on a tRNA corresponds to an anticodon on the tRNA which corresponds to a codon in an mRNA molecule. This rigid system allows for accurate and consistent mRNA translation and protein production. An aaRS attaches an amino acid onto its corresponding tRNA by catalyzing the transesterification of a specific cognate amino acid or its precursor to one of its compatible cognate tRNAs to form an aminoacyl-tRNA. In humans, for example, the 20 different types of aminoacyl-tRNAs are made by the 20 different aaRSs, one for each amino acid of the genetic code.

[0068] An aaRS functions by first binding adenosine triphosphate (ATP) and the corresponding amino acid, or its precursor, to form an aminoacyl-adenylate and releasing inorganic pyrophosphate. The adenylate- aaRS complex then binds the appropriate tRNA’s D arm, and the amino acid is transferred to either the 2’- or 3 ’-OH of the last tRNA nucleotide at the 3 ’-end. Each aaRS is paired with a tRNA (aaRS-tRNA) to ensure accurate and consistent mRNA translation and protein production.

[0069] The present disclosure relates, in part, to aaRS-tRNA pairs that function in (e.g., are endogenous to) a first organism and can be expressed in a cell of or cell lysate derived from a second (e.g., different / heterologous) organism (FIG. 1). In some embodiments, the aaRS- tRNA pairs derived from a first organism are heterologous to a second organism. The term “heterologous,” as used herein, refers to an aaRS, tRNA, or aaRS-tRNA pair that is derived from a different organism. In some embodiments, the aaRS-tRNA pairs derived from a first organism are orthogonal to a second organism. The term “orthogonal,” as used herein, refers to an aaRS-tRNA pair that functions independently. The present disclosure relates, in part, to the discovery of aaRS-tRNA pairs derived from a first organism that can produce a protein of interest from a nucleic acid encoding the protein of interest in a cell of or lysate derived from a different organism. Importantly, tRNA molecules of the present disclosure are only charged by their cognate aaRSs and not by aaRSs present in organisms other than the organism from which the aaRS-tRNA pair was derived. The tRNA molecules of the present disclosure can be used in an organism different from the organism from which the tRNA molecules were derived, and the tRNA molecules are only charged by the aaRSs derived from the same organism as the tRNA molecules and not by aaRSs from different organisms. In some embodiments, heterologous and orthogonal aaRS-tRNA pairs of the present disclosure are specific to each other. In some embodiments, heterologous and orthogonal tRNA, expressed in a cell of or cell lysate derived from a different organism, is only charged in the cell of or cell lysate derived from the different organism if the cognate heterologous and orthogonal aaRS is also present in the cell of or cell lysate derived from the different organism. In some embodiments, fewer than 20%, 15%, 10%, 5%, 2.5%, or 1% of heterologous and orthogonal tRNA molecules are charged by aaRSs other than their cognate aaRSs. In some embodiments, a heterologous and orthogonal tRNA molecule is only charged by its cognate aaRS.

[0070] Aspects of the present disclosure relate to orthogonal aaRS-tRNA pairs and / or the identification of such aaRS-tRNA pairs. In some embodiments, an orthogonal tRNA is derived from Arabidopsis thaliana (A. thaliana), Caenorhabditis elegans (C. elegans), Dictyostelium discoideum (D. discoideum). Drosophila melanogaster (D. melanogaster). Encephalitozoon cuniculi (E. cuniculi), Gallus gallus G. gallus), Lactobacillus johnsonii (L. johnsonii). Candidatus Nitrosocosmicus oleophilus (C. nitrosocosmicus oleophilus). Halomicrobium muhohataei DSM 12286 (H. mukohataei DSM 12286), or Methanoculleus bourgensis (M. bourgensis). In some embodiments, an orthogonal aaRS is derived from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilus, H. mukohataei DSM 12286, or M. bourgensis. In some embodiments, an orthogonal aaRS-tRNA pair is derived from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilus, H. mukohataei DSM 12286, or M. bourgensis. In some embodiments, an orthogonal aaRS-tRNA pair is derived from A. thaliana. In some embodiments, an orthogonal aaRS-tRNA pair is derived from C. elegans. In some embodiments, an orthogonal aaRS- tRNA pair is derived from D. discoideum. In some embodiments, an orthogonal aaRS-tRNA pair is derived from D. melanogaster. In some embodiments, an orthogonal aaRS-tRNA pair is derived from E. cuniculi. In some embodiments, an orthogonal aaRS-tRNA pair is derived from G. gallus. In some embodiments, an orthogonal aaRS-tRNA pair is derived from L. johnsonii. In some embodiments, an orthogonal aaRS-tRNA pair is derived from C. nitrosocosmicus oleophilus. In some embodiments, an orthogonal aaRS-tRNA pair is derived from H. mukohataei DSM 12286. In some embodiments, an orthogonal aaRS-tRNA pair is derived from M. bourgensis.

[0071] In some embodiments, an orthogonal tRNA comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or at least 99.5% identity to any one of SEQ ID NOs: 1-727. In some embodiments, an orthogonal tRNA comprises an amino acid sequences of any one of SEQ ID NOs: 1-727. In some embodiments, an orthogonal aaRS comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or at least 99.5% identity to any one of SEQ ID NOs: 728-1454. In some embodiments, an orthogonal aaRS comprises an amino acid sequences of any one of SEQ ID NOs: 728-1454.

[0072] In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 728. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 2 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 729. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 3 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 730. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 4 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 731. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 5 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 732. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 6 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 733. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 7 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 734. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 8 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 735. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 9 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 736. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 10 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 737. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 11 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 738. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 12 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 739. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 13 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 740. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 14 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 741. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 15 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 742. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 16 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 743. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 17 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 744. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 18 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 745. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 19 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 746. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 20 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 747. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 21 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 748. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 22 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 749. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 23 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 750. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 24 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 751. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 25 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 752. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 26 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 753. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 27 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 754. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 28 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 755. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 29 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 756. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 30 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 757. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 31 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 758. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 32 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 759. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 33 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 760. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 34 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 761. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 35 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 762. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 36 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 763. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 37 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 764. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 38 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 765. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 39 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 766. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 40 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 767. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 41 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 768. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 42 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 769. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 43 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 770. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 44 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 771. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 45 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 772. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 46 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 773. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 47 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 774. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 48 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 775. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 49 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 776. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 50 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 777. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 51 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 778. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 52 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 779. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 53 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 780. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 54 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 781. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 55 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 782. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 56 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 783. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 57 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 784. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 58 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 785. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 59 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 786. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 60 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 787. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 61 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 788. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 62 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 789. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 63 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 790. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 64 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 791. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 65 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 792. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 66 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 793. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 67 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 794. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 68 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 795. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 69 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 796. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 70 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 797. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 71 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 798. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 72 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 799. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 73 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 800. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 74 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 801. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 75 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 802. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 76 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 803. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 77 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 804. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 78 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 805. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 79 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 806. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 80 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 807. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 81 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 808. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 82 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 809. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 83 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 810. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 84 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 811. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 85 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 812. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 86 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 813. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 87 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 814. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 88 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 815. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 89 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 816. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 90 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 817. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 91 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 818. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 92 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 819. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 93 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 820. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 94 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 821. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 95 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 822. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 96 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 823. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 97 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 824. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 98 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 825. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 99 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 826. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 100 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 827. In some embodiments, an orthogonal aaRS- tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 101 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 828. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 102 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 829. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 103 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 830. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 104 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 831. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 105 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 832. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 106 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 833. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 107 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 834. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 108 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 835. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 109 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 836. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 110 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 837. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 111 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 838. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 112 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 839. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 113 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 840. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 114 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 841. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 115 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 842. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 116 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 843. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 117 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 844. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 118 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 845. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 119 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 846. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 120 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 847. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 121 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 848. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 122 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 849. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 123 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 850. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 124 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 851. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 125 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 852. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 126 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 853. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 127 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 854. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 128 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 855. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 129 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 856. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 130 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 857. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 131 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 858. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 132 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 859. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 133 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 860. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 134 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 861. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 135 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 862. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 136 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) 1 identity to SEQ ID NO: 863. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 137 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 864. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 138 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 865. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 139 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 866. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 140 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 867. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 141 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 868. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 142 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 869. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 143 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 870. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 144 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 871. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 145 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 872. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 146 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 873. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 147 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 874. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 148 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 875. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 149 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 876. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 150 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 877. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 151 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 878. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 152 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 879. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 153 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 880. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 154 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 881. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 155 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 882. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 156 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 883. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 157 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 884. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 158 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 885. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 159 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 886. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 160 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 887. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 161 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 888. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 162 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 889. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 163 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 890. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 164 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 891. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 165 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 892. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 166 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 893. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 167 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 894. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 168 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 895. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 169 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 896. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 170 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 897. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 171 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 898. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 172 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 899. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 173 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 900. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 174 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 901. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 175 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 902. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 176 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 903. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 177 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 904. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 178 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 905. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 179 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 906. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 180 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 907. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 181 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 908. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 182 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 909. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 183 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 910. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 184 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 911. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 185 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 912. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 186 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 913. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 187 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 914. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 188 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 915. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 189 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 916. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 190 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 917. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 191 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 918. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 192 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 919. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 193 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 920. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 194 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 921. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 195 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 922. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 196 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 923. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 197 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 924. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 198 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 925. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 199 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 926. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 200 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 927. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 201 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 928. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 202 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 929. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 203 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 930. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 204 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 931. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 205 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 932. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 206 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 933. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 207 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 934. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 208 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 935. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 209 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 936. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 210 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 937. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 211 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 938. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 212 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 939. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 213 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 940. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 214 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 941. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 215 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 942. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 216 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 943. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 217 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 944. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 218 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 945. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 219 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 946. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 220 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 947. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 221 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 948. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 222 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 949. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 223 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 950. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 224 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 951. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 225 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 952. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 226 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 953. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 227 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 954. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 228 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 955. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 229 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 956. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 230 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 957. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 231 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 958. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 232 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 959. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 233 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 960. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 234 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 961. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 235 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 962. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 236 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 963. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 237 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 964. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 238 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 965. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 239 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 966. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 240 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 967. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 241 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 968. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 242 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 969. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 243 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 970. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 244 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 971. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 245 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 972. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 246 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 973. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 247 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 974. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 248 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 975. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 249 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 976. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 250 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 977. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 251 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 978. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 252 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 979. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 253 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 980. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 254 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 981. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 255 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 982. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 256 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 983. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 257 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 984. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 258 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 985. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 259 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 986. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 260 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 987. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 261 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 988. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 262 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 989. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 263 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 990. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 264 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 991. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 265 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 992. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 266 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 993. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 267 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 994. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 268 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 995. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 269 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 996. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 270 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 997. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 271 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 998. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 272 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 999. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 273 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1000. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 274 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1001. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 275 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1002. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 276 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1003. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 277 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1004. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 278 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1005. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 279 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1006. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 280 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1007. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 281 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1008. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 282 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1009. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 283 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1010. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 284 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1011. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 285 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1012. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 286 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1013. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 287 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1014. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 288 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1015. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 289 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1016. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 290 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1017. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 291 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1018. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 292 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1019. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 293 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1020. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 294 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1021. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 295 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1022. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 296 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1023. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 297 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1024. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 298 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1025. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 299 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1026. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 300 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1027. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 301 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1028. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 302 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1029. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 303 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1030. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 304 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1031. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 305 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1032. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 306 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1033. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 307 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1034. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 308 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1035. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 309 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1036. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 310 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1037. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 311 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1038. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 312 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1039. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 313 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1040. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 314 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1041. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 315 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1042. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 316 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1043. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 317 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1044. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 318 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1045. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 319 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1046. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 320 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1047. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 321 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1048. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 322 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1049. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 323 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1050. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 324 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1051. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 325 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1052. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 326 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1053. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 327 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1054. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 328 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1055. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 329 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1056. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 330 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1057. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 331 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1058. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 332 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1059. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 333 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1060. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 334 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1061. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 335 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1062. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 336 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1063. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 337 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1064. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 338 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1065. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 339 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1066. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 340 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1067. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 341 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1068. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 342 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1069. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 343 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1070. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 344 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1071. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 345 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1072. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 346 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1073. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 347 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1074. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 348 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1075. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 349 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1076. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 350 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1077. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 351 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1078. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 352 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1079. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 353 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1080. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 354 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1081. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 355 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1082. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 356 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1083. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 357 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1084. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 358 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1085. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 359 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1086. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 360 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1087. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 361 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1088. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 362 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1089. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 363 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1090. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 364 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1091. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 365 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1092. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 366 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1093. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 367 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1094. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 368 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1095. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 369 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1096. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 370 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1097. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 371 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1098. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 372 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1099. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 373 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1100. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 374 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1101. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 375 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1102. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 376 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1103. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 377 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1104. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 378 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1105. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 379 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1106. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 380 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1107. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 381 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1108. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 382 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1109. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 383 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1110. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 384 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1111. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 385 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1112. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 386 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1113. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 387 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1114. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 388 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1115. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 389 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1116. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 390 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1117. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 391 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1118. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 392 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1119. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 393 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1120. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 394 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1121. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 395 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1122. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 396 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1123. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 397 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1124. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 398 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1125. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 399 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1126. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 400 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1127. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 401 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1128. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 402 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1129. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 403 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1130. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 404 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1131. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 405 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1132. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 406 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1133. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 407 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1134. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 408 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1135. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 409 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1136. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 410 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1137. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 411 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1138. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 412 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1139. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 413 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1140. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 414 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1141. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 415 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1142. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 416 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1143. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 417 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1144. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 418 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1145. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 419 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1146. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 420 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1147. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 421 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1148. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 422 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1149. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 423 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1150. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 424 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1151. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 425 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1152. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 426 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1153. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 427 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1154. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 428 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1155. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 429 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1156. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 430 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1157. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 431 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1158. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 432 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1159. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 433 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1160. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 434 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1161. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 435 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1162. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 436 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1163. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 437 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1164. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 438 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1165. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 439 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1166. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 440 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1167. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 441 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1168. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 442 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1169. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 443 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1170. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 444 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1171. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 445 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1172. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 446 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1173. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 447 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1174. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 448 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1175. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 449 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1176. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 450 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1177. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 451 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1178. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 452 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1179. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 453 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1180. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 454 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1181. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 455 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1182. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 456 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1183. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 457 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1184. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 458 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1185. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 459 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1186. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 460 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1187. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 461 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1188. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 462 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1189. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 463 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1190. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 464 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1191. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 465 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1192. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 466 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1193. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 467 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1194. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 468 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1195. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 469 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1196. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 470 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1197. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 471 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1198. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 472 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1199. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 473 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1200. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 474 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1201. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 475 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1202. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 476 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1203. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 477 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1204. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 478 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1205. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 479 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1206. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 480 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1207. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 481 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1208. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 482 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1209. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 483 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1210. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 484 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1211. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 485 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1212. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 486 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1213. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 487 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1214. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 488 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1215. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 489 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1216. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 490 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1217. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 491 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1218. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 492 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1219. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 493 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1220. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 494 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1221. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 495 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1222. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 496 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1223. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 497 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1224. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 498 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1225. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 499 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1226. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 500 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1227. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 501 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1228. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 502 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1229. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 503 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1230. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 504 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1231. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 505 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1232. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 506 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1233. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 507 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1234. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 508 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1235. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 509 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1236. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 510 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1237. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 511 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1238. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 512 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1239. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 513 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1240. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 514 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1241. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 515 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1242. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 516 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1243. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 517 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1244. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 518 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1245. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 519 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1246. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 520 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1247. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 521 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1248. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 522 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1249. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 523 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1250. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 524 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1251. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 525 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1252. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 526 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1253. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 527 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1254. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 528 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1255. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 529 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1256. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 530 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1257. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 531 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1258. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 532 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1259. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 533 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1260. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 534 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1261. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 535 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1262. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 536 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1263. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 537 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1264. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 538 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1265. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 539 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1266. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 540 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1267. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 541 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1268. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 542 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1269. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 543 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1270. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 544 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1271. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 545 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1272. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 546 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1273. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 547 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1274. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 548 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1275. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 549 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1276. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 550 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1277. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 551 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1278. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 552 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1279. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 553 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1280. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 554 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1281. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 555 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1282. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 556 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1283. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 557 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1284. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 558 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1285. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 559 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1286. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 560 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1287. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 561 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1288. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 562 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1289. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 563 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1290. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 564 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1291. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 565 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1292. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 566 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1293. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 567 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1294. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 568 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1295. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 569 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1296. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 570 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1297. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 571 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1298. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 572 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1299. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 573 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1300. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 574 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1301. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 575 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1302. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 576 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1303. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 577 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1304. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 578 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1305. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 579 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1306. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 580 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1307. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 581 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1308. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 582 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1309. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 583 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1310. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 584 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1311. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 585 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1312. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 586 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1313. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 587 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1314. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 588 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1315. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 589 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1316. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 590 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1317. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 591 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1318. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 592 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1319. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 593 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1320. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 594 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1321. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 595 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1322. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 596 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1323. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 597 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1324. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 598 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1325. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 599 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1326. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 600 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1327. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 601 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1328. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 602 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1329. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 603 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1330. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 604 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1331. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 605 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1332. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 606 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1333. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 607 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1334. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 608 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1335. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 609 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1336. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 610 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1337. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 611 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1338. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 612 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1339. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 613 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1340. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 614 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1341. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 615 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1342. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 616 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1343. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 617 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1344. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 618 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1345. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 619 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1346. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 620 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1347. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 621 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1348. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 622 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1349. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 623 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1350. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 624 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1351. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 625 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1352. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 626 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1353. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 627 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1354. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 628 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1355. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 629 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1356. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 630 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1357. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 631 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1358. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 632 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1359. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 633 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1360. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 634 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1361. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 635 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1362. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 636 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1363. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 637 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1364. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 638 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1365. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 639 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1366. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 640 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1367. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 641 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1368. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 642 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1369. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 643 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1370. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 644 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1371. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 645 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1372. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 646 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1373. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 647 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1374. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 648 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1375. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 649 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1376. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 650 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1377. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 651 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1378. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 652 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1379. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 653 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1380. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 654 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1381. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 655 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1382. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 656 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1383. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 657 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1384. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 658 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1385. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 659 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1386. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 660 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1387. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 661 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1388. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 662 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1389. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 663 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1390. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 664 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1391. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 665 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1392. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 666 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1393. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 667 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1394. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 668 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1395. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 669 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1396. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 670 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1397. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 671 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1398. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 672 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1399. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 673 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1400. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 674 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1401. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 675 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1402. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 676 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1403. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 677 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1404. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 678 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1405. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 679 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1406. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 680 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1407. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 681 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1408. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 682 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1409. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 683 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1410. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 684 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1411. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 685 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1412. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 686 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1413. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 687 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1414. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 688 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1415. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 689 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1416. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 690 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1417. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 691 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1418. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 692 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1419. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 693 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1420. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 694 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1421. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 695 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1422. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 696 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1423. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 697 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1424. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 698 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1425. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 699 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1426. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 700 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1427. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 701 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1428. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 702 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1429. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 703 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1430. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 704 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1431. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 705 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1432. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 706 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1433. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 707 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1434. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 708 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1435. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 709 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1436. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 710 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1437. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 711 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1438. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 712 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1439. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 713 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1440. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 714 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1441. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 715 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1442. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 716 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1443. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 717 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1444. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 718 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1445. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 719 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1446. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 720 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1447. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 721 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1448. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 722 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1449. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 723 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1450. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 724 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1451. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 725 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1452. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 726 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1453. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 727 and an aaRS comprising a sequence having at least 80% (e.g., at least 85%, at least 90%, or at least 95%) identity to SEQ ID NO: 1454. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 1 and an aaRS comprising the sequence of SEQ ID NO: 728. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 2 and an aaRS comprising the sequence of SEQ ID NO: 729. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 3 and an aaRS comprising the sequence of SEQ ID NO: 730. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 4 and an aaRS comprising the sequence of SEQ ID NO: 731. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 5 and an aaRS comprising the sequence of SEQ ID NO: 732. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 6 and an aaRS comprising the sequence of SEQ ID NO: 733. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 7 and an aaRS comprising the sequence of SEQ ID NO: 734. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 8 and an aaRS comprising the sequence of SEQ ID NO: 735. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 9 and an aaRS comprising the sequence of SEQ ID NO: 736. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 10 and an aaRS comprising the sequence of SEQ ID NO: 737. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 11 and an aaRS comprising the sequence of SEQ ID NO: 738. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 12 and an aaRS comprising the sequence of SEQ ID NO: 739. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 13 and an aaRS comprising the sequence of SEQ ID NO: 740. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 14 and an aaRS comprising the sequence of SEQ ID NO: 741. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 15 and an aaRS comprising the sequence of SEQ ID NO: 742. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 16 and an aaRS comprising the sequence of SEQ ID NO: 743. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 17 and an aaRS comprising the sequence of SEQ ID NO: 744. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 18 and an aaRS comprising the sequence of SEQ ID NO: 745. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 19 and an aaRS comprising the sequence of SEQ ID NO: 746. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 20 and an aaRS comprising the sequence of SEQ ID NO: 747. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 21 and an aaRS comprising the sequence of SEQ ID NO: 748. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 22 and an aaRS comprising the sequence of SEQ ID NO: 749. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 23 and an aaRS comprising the sequence of SEQ ID NO: 750. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 24 and an aaRS comprising the sequence of SEQ ID NO: 751. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 25 and an aaRS comprising the sequence of SEQ ID NO: 752. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 26 and an aaRS comprising the sequence of SEQ ID NO: 753. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 27 and an aaRS comprising the sequence of SEQ ID NO: 754. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 28 and an aaRS comprising the sequence of SEQ ID NO: 755. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 29 and an aaRS comprising the sequence of SEQ ID NO: 756. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 30 and an aaRS comprising the sequence of SEQ ID NO: 757. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 31 and an aaRS comprising the sequence of SEQ ID NO: 758. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 32 and an aaRS comprising the sequence of SEQ ID NO: 759. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 33 and an aaRS comprising the sequence of SEQ ID NO: 760. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 34 and an aaRS comprising the sequence of SEQ ID NO: 761. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 35 and an aaRS comprising the sequence of SEQ ID NO: 762. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 36 and an aaRS comprising the sequence of SEQ ID NO: 763. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 37 and an aaRS comprising the sequence of SEQ ID NO: 764. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 38 and an aaRS comprising the sequence of SEQ ID NO: 765. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 39 and an aaRS comprising the sequence of SEQ ID NO: 766. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 40 and an aaRS comprising the sequence of SEQ ID NO: 767. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 41 and an aaRS comprising the sequence of SEQ ID NO: 768. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 42 and an aaRS comprising the sequence of SEQ ID NO: 769. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 43 and an aaRS comprising the sequence of SEQ ID NO: 770. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 44 and an aaRS comprising the sequence of SEQ ID NO: 771. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 45 and an aaRS comprising the sequence of SEQ ID NO: 772. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 46 and an aaRS comprising the sequence of SEQ ID NO: 773. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 47 and an aaRS comprising the sequence of SEQ ID NO: 774. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 48 and an aaRS comprising the sequence of SEQ ID NO: 775. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 49 and an aaRS comprising the sequence of SEQ ID NO: 776. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 50 and an aaRS comprising the sequence of SEQ ID NO: 777. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 51 and an aaRS comprising the sequence of SEQ ID NO: 778. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 52 and an aaRS comprising the sequence of SEQ ID NO: 779. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 53 and an aaRS comprising the sequence of SEQ ID NO: 780. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 54 and an aaRS comprising the sequence of SEQ ID NO: 781. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 55 and an aaRS comprising the sequence of SEQ ID NO: 782. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 56 and an aaRS comprising the sequence of SEQ ID NO: 783. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 57 and an aaRS comprising the sequence of SEQ ID NO: 784. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 58 and an aaRS comprising the sequence of SEQ ID NO: 785. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 59 and an aaRS comprising the sequence of SEQ ID NO: 786. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 60 and an aaRS comprising the sequence of SEQ ID NO: 787. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 61 and an aaRS comprising the sequence of SEQ ID NO: 788. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 62 and an aaRS comprising the sequence of SEQ ID NO: 789. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 63 and an aaRS comprising the sequence of SEQ ID NO: 790. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 64 and an aaRS comprising the sequence of SEQ ID NO: 791. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 65 and an aaRS comprising the sequence of SEQ ID NO: 792. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 66 and an aaRS comprising the sequence of SEQ ID NO: 793. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 67 and an aaRS comprising the sequence of SEQ ID NO: 794. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 68 and an aaRS comprising the sequence of SEQ ID NO: 795. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 69 and an aaRS comprising the sequence of SEQ ID NO: 796. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 70 and an aaRS comprising the sequence of SEQ ID NO: 797. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 71 and an aaRS comprising the sequence of SEQ ID NO: 798. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 72 and an aaRS comprising the sequence of SEQ ID NO: 799. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 73 and an aaRS comprising the sequence of SEQ ID NO: 800. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 74 and an aaRS comprising the sequence of SEQ ID NO: 801. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 75 and an aaRS comprising the sequence of SEQ ID NO: 802. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 76 and an aaRS comprising the sequence of SEQ ID NO: 803. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 77 and an aaRS comprising the sequence of SEQ ID NO: 804. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 78 and an aaRS comprising the sequence of SEQ ID NO: 805. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 79 and an aaRS comprising the sequence of SEQ ID NO: 806. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 80 and an aaRS comprising the sequence of SEQ ID NO: 807. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 81 and an aaRS comprising the sequence of SEQ ID NO: 808. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 82 and an aaRS comprising the sequence of SEQ ID NO: 809. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 83 and an aaRS comprising the sequence of SEQ ID NO: 810. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 84 and an aaRS comprising the sequence of SEQ ID NO: 811. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 85 and an aaRS comprising the sequence of SEQ ID NO: 812. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 86 and an aaRS comprising the sequence of SEQ ID NO: 813. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 87 and an aaRS comprising the sequence of SEQ ID NO: 814. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 88 and an aaRS comprising the sequence of SEQ ID NO: 815. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 89 and an aaRS comprising the sequence of SEQ ID NO: 816. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 90 and an aaRS comprising the sequence of SEQ ID NO: 817. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 91 and an aaRS comprising the sequence of SEQ ID NO: 818. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 92 and an aaRS comprising the sequence of SEQ ID NO: 819. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 93 and an aaRS comprising the sequence of SEQ ID NO: 820. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 94 and an aaRS comprising the sequence of SEQ ID NO: 821. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 95 and an aaRS comprising the sequence of SEQ ID NO: 822. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 96 and an aaRS comprising the sequence of SEQ ID NO: 823. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 97 and an aaRS comprising the sequence of SEQ ID NO: 824. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 98 and an aaRS comprising the sequence of SEQ ID NO: 825. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 99 and an aaRS comprising the sequence of SEQ ID NO: 826. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 100 and an aaRS comprising the sequence of SEQ ID NO: 827. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 101 and an aaRS comprising the sequence of SEQ ID NO: 828. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 102 and an aaRS comprising the sequence of SEQ ID NO: 829. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 103 and an aaRS comprising the sequence of SEQ ID NO: 830. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 104 and an aaRS comprising the sequence of SEQ ID NO: 831. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 105 and an aaRS comprising the sequence of SEQ ID NO: 832. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 106 and an aaRS comprising the sequence of SEQ ID NO: 833. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 107 and an aaRS comprising the sequence of SEQ ID NO: 834. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 108 and an aaRS comprising the sequence of SEQ ID NO: 835. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 109 and an aaRS comprising the sequence of SEQ ID NO: 836. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 110 and an aaRS comprising the sequence of SEQ ID NO: 837. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 111 and an aaRS comprising the sequence of SEQ ID NO: 838. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 112 and an aaRS comprising the sequence of SEQ ID NO: 839. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 113 and an aaRS comprising the sequence of SEQ ID NO: 840. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 114 and an aaRS comprising the sequence of SEQ ID NO: 841. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 115 and an aaRS comprising the sequence of SEQ ID NO: 842. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 116 and an aaRS comprising the sequence of SEQ ID NO: 843. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 117 and an aaRS comprising the sequence of SEQ ID NO: 844. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 118 and an aaRS comprising the sequence of SEQ ID NO: 845. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 119 and an aaRS comprising the sequence of SEQ ID NO: 846. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 120 and an aaRS comprising the sequence of SEQ ID NO: 847. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 121 and an aaRS comprising the sequence of SEQ ID NO: 848. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 122 and an aaRS comprising the sequence of SEQ ID NO: 849. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 123 and an aaRS comprising the sequence of SEQ ID NO: 850. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 124 and an aaRS comprising the sequence of SEQ ID NO: 851. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 125 and an aaRS comprising the sequence of SEQ ID NO: 852. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 126 and an aaRS comprising the sequence of SEQ ID NO: 853. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 127 and an aaRS comprising the sequence of SEQ ID NO: 854. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 128 and an aaRS comprising the sequence of SEQ ID NO: 855. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 129 and an aaRS comprising the sequence of SEQ ID NO: 856. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 130 and an aaRS comprising the sequence of SEQ ID NO: 857. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 131 and an aaRS comprising the sequence of SEQ ID NO: 858. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 132 and an aaRS comprising the sequence of SEQ ID NO: 859. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 133 and an aaRS comprising the sequence of SEQ ID NO: 860. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 134 and an aaRS comprising the sequence of SEQ ID NO: 861. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 135 and an aaRS comprising the sequence of SEQ ID NO: 862. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 136 and an aaRS comprising the sequence of SEQ ID NO: 863. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 137 and an aaRS comprising the sequence of SEQ ID NO: 864. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 138 and an aaRS comprising the sequence of SEQ ID NO: 865. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 139 and an aaRS comprising the sequence of SEQ ID NO: 866. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 140 and an aaRS comprising the sequence of SEQ ID NO: 867. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 141 and an aaRS comprising the sequence of SEQ ID NO: 868. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 142 and an aaRS comprising the sequence of SEQ ID NO: 869. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 143 and an aaRS comprising the sequence of SEQ ID NO: 870. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 144 and an aaRS comprising the sequence of SEQ ID NO: 871. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 145 and an aaRS comprising the sequence of SEQ ID NO: 872. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 146 and an aaRS comprising the sequence of SEQ ID NO: 873. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 147 and an aaRS comprising the sequence of SEQ ID NO: 874. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 148 and an aaRS comprising the sequence of SEQ ID NO: 875. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 149 and an aaRS comprising the sequence of SEQ ID NO: 876. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 150 and an aaRS comprising the sequence of SEQ ID NO: 877. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 151 and an aaRS comprising the sequence of SEQ ID NO: 878. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 152 and an aaRS comprising the sequence of SEQ ID NO: 879. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 153 and an aaRS comprising the sequence of SEQ ID NO: 880. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 154 and an aaRS comprising the sequence of SEQ ID NO: 881. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 155 and an aaRS comprising the sequence of SEQ ID NO: 882. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 156 and an aaRS comprising the sequence of SEQ ID NO: 883. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 157 and an aaRS comprising the sequence of SEQ ID NO: 884. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 158 and an aaRS comprising the sequence of SEQ ID NO: 885. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 159 and an aaRS comprising the sequence of SEQ ID NO: 886. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 160 and an aaRS comprising the sequence of SEQ ID NO: 887. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 161 and an aaRS comprising the sequence of SEQ ID NO: 888. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 162 and an aaRS comprising the sequence of SEQ ID NO: 889. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 163 and an aaRS comprising the sequence of SEQ ID NO: 890. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 164 and an aaRS comprising the sequence of SEQ ID NO: 891. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 165 and an aaRS comprising the sequence of SEQ ID NO: 892. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 166 and an aaRS comprising the sequence of SEQ ID NO: 893. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 167 and an aaRS comprising the sequence of SEQ ID NO: 894. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 168 and an aaRS comprising the sequence of SEQ ID NO: 895. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 169 and an aaRS comprising the sequence of SEQ ID NO: 896. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 170 and an aaRS comprising the sequence of SEQ ID NO: 897. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 171 and an aaRS comprising the sequence of SEQ ID NO: 898. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 172 and an aaRS comprising the sequence of SEQ ID NO: 899. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 173 and an aaRS comprising the sequence of SEQ ID NO: 900. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 174 and an aaRS comprising the sequence of SEQ ID NO: 901. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 175 and an aaRS comprising the sequence of SEQ ID NO: 902. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 176 and an aaRS comprising the sequence of SEQ ID NO: 903. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 177 and an aaRS comprising the sequence of SEQ ID NO: 904. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 178 and an aaRS comprising the sequence of SEQ ID NO: 905. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 179 and an aaRS comprising the sequence of SEQ ID NO: 906. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 180 and an aaRS comprising the sequence of SEQ ID NO: 907. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 181 and an aaRS comprising the sequence of SEQ ID NO: 908. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 182 and an aaRS comprising the sequence of SEQ ID NO: 909. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 183 and an aaRS comprising the sequence of SEQ ID NO: 910. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 184 and an aaRS comprising the sequence of SEQ ID NO: 911. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 185 and an aaRS comprising the sequence of SEQ ID NO: 912. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 186 and an aaRS comprising the sequence of SEQ ID NO: 913. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 187 and an aaRS comprising the sequence of SEQ ID NO: 914. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 188 and an aaRS comprising the sequence of SEQ ID NO: 915. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 189 and an aaRS comprising the sequence of SEQ ID NO: 916. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 190 and an aaRS comprising the sequence of SEQ ID NO: 917. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 191 and an aaRS comprising the sequence of SEQ ID NO: 918. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 192 and an aaRS comprising the sequence of SEQ ID NO: 919. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 193 and an aaRS comprising the sequence of SEQ ID NO: 920. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 194 and an aaRS comprising the sequence of SEQ ID NO: 921. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 195 and an aaRS comprising the sequence of SEQ ID NO: 922. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 196 and an aaRS comprising the sequence of SEQ ID NO: 923. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 197 and an aaRS comprising the sequence of SEQ ID NO: 924. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 198 and an aaRS comprising the sequence of SEQ ID NO: 925. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising the sequence of SEQ ID NO: 199 and an aaRS comprising the sequence of SEQ ID NO: 926. In some embodiments, an orthogonal aaRS-tRNA pair comprises a tRNA comprising ...

Claims

CLAIMSWhat is claimed is:

1. A host cell comprising: a transfer ribonucleic acid (tRNA); and an aminoacyl-tRNA synthetase (aaRS); wherein the tRNA and aaRS are from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilius, H. mukohataei DSM 12286, or M. bourgensis, and the tRNA and aaRS are heterologous to the host cell.

2. The host cell of claim 1, wherein the tRNA comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 1-727.

3. The host cell of claim 1 or 2, wherein the aaRS comprises an amino acid sequence having at least 80% identity to any one of SEQ ID NOs: 728-1454.

4. The host cell of any one of claims 1-3, wherein the tRNA comprises an amino acid sequence having at least 90% or at least 95% identity to any one of SEQ ID NOs: 1-727 and the aaRS comprises an amino acid sequence having at least 90% or at least 95% identity to any one of SEQ ID NOs: 728-1454.

5. A host cell comprising: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728- 1454; wherein the tRNA and aaRS are heterologous to host cell.

6. A host cell comprising: a transfer ribonucleic acid (tRNA) comprising an amino acid sequence having 80%- 99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454, optionally wherein the tRNA and aaRS are heterologous to host cell.

7. The host cell of any one of claims 1-6, wherein the tRNA is charged with a standard amino acid.

8. The host cell of any one of claims 1-6, wherein the tRNA is charged with a nonstandard amino acid.

9. The host cell of claim 8, wherein the nonstandard amino acid is cystine, desmosine, isodesmosine, hydroxyproline, hydroxylysine, gamma-carboxyglutamate, phosphoserine, phosphothreonine, phosphotyrosine, N-acetyl lysine, methyllysine, para-azido-phenylalanine, propargyloxy -phenylalanine, benzoyl-phenylalanine, or dibenzocyclooctane-lysine. ..

10. The host cell of any one of claim 1-9, wherein the host cell is a prokaryotic cell.

11. The host cell of claim 10, wherein the prokaryotic cell is a bacterial cell or an archaebacterial cell.

12. The host cell of claim 11, wherein the bacterial cell is an Escherichia coli (E. coli) cell.

13. The host cell of any one of claim 1-9, wherein the host cell is a eukaryotic cell.

14. The host cell of claim 13, wherein the eukaryotic cell is an animal cell, an insect cell, a fungal cell, or an algal cell.

15. The host cell of claim 14, wherein the animal cell is a mammalian cell.

16. The host cell of claim 15, wherein the mammalian cell is a human cell.

17. The host cell of claim 14, wherein the insect cell is an Sf9 cell, a Tni cell, an Sf21 cell, or an S2 cell.

18. A cell-free system, comprising: a transfer ribonucleic acid (tRNA); andan aminoacyl-tRNA synthetase (aaRS); wherein the tRNA and aaRS are from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilius, H. mukohataei DSM 12286, or M. bourgensis, and the tRNA and aaRS are heterologous to the cell-free system.

19. A cell-free system comprising: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728- 1454; wherein the tRNA and aaRS are heterologous to cell-free system.

20. A cell-free system comprising: a transfer ribonucleic acid (tRNA) comprising an amino acid sequence having 80%- 99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454, optionally wherein the tRNA and aaRS are heterologous to cell-free system.

21. A composition comprising: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1-727; an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728-1454; and a nonstandard amino acid.

22. A composition comprising: a transfer ribonucleic acid (tRNA) comprising an amino acid sequence having 80%- 99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1-727; an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454; and optionally a nonstandard amino acid.

23. A method of producing a protein of interest, comprising: delivering to a host cell a transfer ribonucleic acid (tRNA) and an aminoacyl-tRNA synthetase (aaRS), wherein the tRNA and aaRS are from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilius, H. mukohataei DSM 12286, or M. bourgensis, and the tRNA and aaRS are heterologous to the host cell; and optionally delivering to the host cell a nucleic acid encoding a protein of interest.

24. A method of producing a protein of interest, comprising:(a) delivering to a host cell: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728-1454, wherein the tRNA and aaRS are heterologous to the host cell; and(b) optionally delivering to the host cell a nucleic acid encoding a protein of interest.

25. A method of producing a protein of interest, comprising:(a) delivering to a host cell: a transfer ribonucleic acid (tRNA) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454, optionally wherein the tRNA and aaRS are heterologous to the host cell; and(b) optionally delivering to the host cell a nucleic acid encoding a protein of interest.

26. A method of producing a protein of interest, comprising: delivering to a lysate a transfer ribonucleic acid (tRNA) and an aminoacyl-tRNA synthetase (aaRS), wherein the tRNA and aaRS are from A. thaliana, C. elegans, D. discoideum, D. melanogaster, E. cuniculi, G. gallus, L. johnsonii, C. nitrosocosmicus oleophilius, H. mukohataei DSM 12286, or M. bourgensis, and the tRNA and aaRS are heterologous to the lysate; andoptionally delivering to the lysate a nucleic acid encoding a protein of interest.

27. A method of producing a protein of interest, comprising:(a) delivering to a lysate: a transfer ribonucleic acid (tRNA) comprising the amino acid sequence of any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising the amino acid sequence of any one of SEQ ID NO: 728-1454, wherein the tRNA and aaRS are heterologous to the lysate; and(b) optionally delivering to the lysate a nucleic acid encoding a protein of interest.

28. A method of producing a protein of interest, comprising:(a) delivering to a lysate: transfer ribonucleic acid (tRNA) comprising an amino acid sequence having80%-99%, 80%-98%, or 80%-95% identity to any one of SEQ ID NO: 1-727; and an aminoacyl-tRNA synthetase (aaRS) comprising an amino acid sequence having 80%-99%, 80%-98%, or 80%-95% identity SEQ ID NO: 728-1454, optionally wherein the tRNA and aaRS are heterologous to the lysate; and(b) optionally delivering to the lysate a nucleic acid encoding a protein of interest.

29. A method of evolving a tRNA-aaRS pair, comprising:(i) evolving a native transfer ribonucleic acid (tRNA) to accept a nonstandard amino acid; and(ii) evolving a native aminoacyl-tRNA synthetase (aaRS) to charge the tRNA with the nonstandard amino acid; the evolved tRNA comprises an amino acid sequence having greater than 75% and less than 100% identity to any one of SEQ ID NOs: 1-727, and wherein the evolved aaRS comprises an amino acid sequence having greater than 75% and less than 100% identity to any one of SEQ ID NOs: 728-1454.

30. The method of claim 29, wherein the native tRNA comprises an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 1-727, and wherein the native aaRS comprises an amino acid sequence that is at least 80% identical to any one of SEQ ID NOs: 728-1454.

31. A method comprising: combining a cell lysate that comprises one or more native aminoacyl tRNA synthetases (aaRSs) with a pool of in vitro transcribed transfer ribonucleic acid (tRNA) molecules; extracting one or more of the synthetic tRNA molecules from the cell lysate to obtain isolated tRNA molecules; and assessing aminoacylation of one or more of the isolated tRNA molecules using periodate oxidation and beta-elimination.

32. A method, comprising: combining:(i) a cell lysate that comprises one or more native aminoacyl tRNA synthetases (aaRSs);(ii) a double- stranded DNA encoding a heterologous aaRS;(iii) a heterologous transfer ribonucleic acid (tRNA); and(iv) a detectably labeled adenosine triphosphate (ATP); and assessing aminoacylation of the tRNA.

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