95 results about "Nucleotide synthesis" patented technology

The present invention relates generally to the fields of oligonucleotide synthesis. More particularly, it concerns the assembly of genes and genomes of completely synthetic artificial organisms. Thus, the present invention outlines a novel approach to utilizing the results of genomic sequence information by computer directed gene synthesis based on computing on the human genome database. Specifically, the present invention contemplates and describes the chemical synthesis and resynthesis of genes defined by the genome sequence in a host vector and transfer and expression of these sequences into suitable hosts.

The present invention relates to oligonucleotide synthesis. In particular, the present invention provides methods for characterizing samples useful for making oligonucleotides.

Compositions and methods are provided for nucleotide analogs comprising protein shields for improving enzyme photostability in single molecule real time sequencing. Nucleotide analogs of the invention have a protein shield between the dye moieties and nucleotide moieties of the analog. The protein prevents the direct interaction of the dye moiety with the enzyme carrying out nucleotide synthesis preventing photodamage to the enzyme. The nucleotide analogs of the invention can have multiple dyes and multiple nucleotide moieties.

Compositions and methods are provided for nucleotide analogs comprising protein shields for improving enzyme photostability in single molecule real time sequencing. Nucleotide analogs of the invention have a protein shield between the dye moieties and nucleotide moieties of the analog. The protein prevents the direct interaction of the dye moiety with the enzyme carrying out nucleotide synthesis preventing photodamage to the enzyme. The nucleotide analogs of the invention can have multiple dyes and multiple nucleotide moieties.

The present invention relates to methods for measuring the proliferation and destruction rates of cells by measuring deoxyribonucleic acid (DNA) synthesis and / or destruction. In particular, the methods utilize non-radioactive stable isotope labels to endogenously label DNA synthesized through the de novo nucleotide synthesis pathway in a cell. The amount of label incorporated in the DNA is measured as an indication of cellular proliferation. The decay of labeled DNA over time is measured as an indication of cellular destruction. Such methods do not involve radioactivity or potentially toxic metabolites, and are suitable for use both in vitro and in vivo. Therefore, the invention is useful for measuring cellular proliferation or cellular destruction rates in humans for the diagnosis, prevention, or management of a variety of disease conditions in which cellular proliferation or cellular destruction is involved. The invention also provides methods for measuring proliferation or destruction of T cells in a subject infected with human immunodeficiency virus (HIV) and methods of screening an agent for a capacity to induce or inhibit cellular proliferation or destruction. In addition, the invention provides methods for measuring cellular proliferation in a proliferating population which utilize both radioactive isotope labels and stable isotopes to endogenously label DNA through the de novo nucleotide synthesis pathway.

Precursors for use in the synthesis of polynucleotides are disclosed. The precursors include a heterocyclic base having an exocyclic amine group and a substituted or unsubstituted triaryl methyl protecting group bound to the exocyclic amine group. In particular embodiments, the precursor has the structure:wherein:O and H represent oxygen and hydrogen, respectively,R1 is hydrido, hydroxyl, protected hydroxyl, lower alkyl, modified lower alkyl, or alkoxy,one of R2 or R3 is a hydroxyl protecting group; and the other of R2 or R3 is a reactive group capable of reacting with a reactive site hydroxyl,Base is a heterocyclic base having an exocyclic amine group, andTram is a triaryl methyl group having the structure (V)wherein the broken line represents a bond to the amino nitrogen of the exocyclic amine group, and R4, R5 and R6 are independently selected from unsubstituted or substituted aryl groups, provided that at least one of R4, R5, and R6 is an aryl group other than phenyl and other than substituted phenyl.

A nucleoside monomer that is protected by a thionocarbamate protecting group and contains one or more 2H, 13C, or 15N isotopes in the ribose and / or base part is provided, as well as a method for making a polynucleotide that uses the same. Also provided is a polynucleotide synthesis method that employs a diamine to deprotect a protected polynucleotide.

The invention provides a method of synthesizing complex mixtures of long polynucleotides by separately synthesizing and assembling shorter component oligonucleotides. In one aspect, pairs of oligonucleotides that form components of such polynucleotides are synthesized on one or more microarrays, or other large-scale parallel solid phase synthesis platforms, after which they are released. Members of each pair contain unique complementary barcode sequences that are used match-up pairs in a hybridization reaction to form duplexes. Such duplexes are then extended with a DNA polymerase and the resulting extension product is amplified to form an amplicon. The amplicon may be either used directly as the desired polynucleotide, or it may undergo further processing, such as capture on solid phase supports and / or additional enzymatic or chemical processing, to produce a desired polynucleotide product, such as a circularizing probe for multiplex analysis of genomic DNA, or the like.

Precursors for use in the synthesis of polynucleotides are disclosed. The precursors include a heterocyclic base having an exocyclic amine group and a substituted or unsubstituted triaryl methyl protecting group bound to the exocyclic amine group.

Precursors for use in the synthesis of polynucleotides and methods of using the precursors in synthesizing polynucleotides are disclosed. The precursors include a heterocyclic base having an exocyclic amine group and a substituted or unsubstituted triaryl methyl protecting group bound to the exocyclic amine group.

The present invention provides methods for detecting the presence of a molecule of interest by generating multiple detectable oligonucleotides through reiterative enzymatic oligonucleotide synthesis events on a polynucleotide sequence and detecting said products. The invention also provides for methods of multiplex abortive transcription. In a further aspect, the invention provides for the use of dendrimers containing abortive promoter cassettes. In one aspect, the invention provides for abortive promoter cassettes suitable for use in the present invention. In one aspect the products of abortive transcription are detected with the use of mass spectrometry. In another aspect, the invention provides a method for detecting a target protein, DNA or RNA by generating multiple detectable RNA oligoribonucleotides by abortive transcription that are detected, including by mass spectrometry.

Methods and systems for automated polynucleotide synthesis design are provided. Example embodiments provide an Automated Polynucleotide Synthesis Design System (“APSDS”), which automatically generates a synthesis design for a designated target sequence specification. In one embodiment, the APSDS comprises a synthesis design engine, user interface support, a synthesis rules data repository, and a synthesis data repository. The APSDS automatically generates a synthesis design by receiving a target sequence(s) specification, generating a potential synthesis design, evaluating the potential design against synthesis rules, and when the evaluation indicates that the potential design is not successful according to the synthesis rules, adjusting the design to generate a new potential synthesis design and repeating the process of evaluating and adjusting until a potential synthesis design is found that satisfies the synthesis rules or until no solution is found.

The invention relates to compositions, methods, and kits for hot start polynucleotide synthesis, including extension of primed polynucleotide templates and polymerase chain reaction (PCR). Hot start is provided by a thermally inactivated blocking polymerase protein that binds primed polynucleotide templates and prevents their access to a thermostable nucleic acid polymerase. High temperatures employed in the synthesis reaction cause the blocking polymerase to denature, thereby permitting the action of a thermostable processive polymerase. Compositions of the invention include a specific blocking polymerase protein which is a mutant of the Klenow fragment of E. coli DNA polymerase. The mutant is essentially devoid of polymerase activity, processivity, and 3′ to 5′ exonuclease activity. Use of the thermally inactivated blocking polymerase together with a thermostable polymerase reduces non-specific priming and accumulation of unwanted amplification products, increasing the specificity and sensitivity of the synthesis reaction.

The present invention discloses a kind of nucleoside phosphoramidite for synthesizing RNA oligonucleotide and its synthesis. The present invention adopts [2, 2-dimethyl-2-(o-nitrophenyl) acetylmethoxy] ether (DNPA) as the 2'-hydroxy protecting radical. The protecting radical DNPA is compatible with convenient 5'-hydroxy protecting radical, and may be eliminated under mild condition, so that it can ensure the RNA integrality and is suitable for synthesis of RNA oligonucleotide. The present invention provides one new path for synthesizing RNA oligonucleotide and may be used for the RNA molecule synthesis of various kinds of natural nucleoside, non-natural nucleoside, labeling nucleoside or their derivatives.

The present invention provides building blocks and methods for synthesizing very pure RNA in a form that can efficiently be modified at the 3′ end. Reverse RNA monomer phosphoramidites have been developed for RNA synthesis in 5′→3′ direction, leading to very clean oligo synthesis that allows for the introduction of various modifications at the 3′-end cleanly and efficiently. Higher coupling efficiency per step have been observed during automated oligo synthesis with the reverse RNA amidites disclosed herein, resulting in a greater ability to achieve higher purity and produce very long oligonucleotides. The use of the reverse RNA phosphoramidites in the synthetic process of this invention leads to oligonucleotides free of N+1 species.

The invention discloses a two-nucleotide synthetic sequencing analysis method for a multi-template PCR product. The method comprises the steps of uniformly dividing the same PCR product into at least two parts, and carrying out two-nucleotide synthetic sequencing, wherein each sequencing reaction obtains sequencing encoding information containing two nucleotide kinds and nucleotide synthetic amount; selecting at least two groups of encoding information respectively obtained by at least two groups of different two-nucleotide synthetic circular sequencing in three groups to decode; and finally, determining the contents of different DNA templates of the PCR product through association analysis. The two-nucleotide synthetic sequencing analysis method can be used for widening the single-nucleotide synthetic sequencing analysis range, directly measuring the proportions of all the DNA templates in a sample and finding and screening nucleotide markers from large-scale samples, and is suitable for sequencing analysis of the multi-template PCR product and a mixed sample PCR product.

The present invention provides methods for detecting the presence of a target molecule by generating multiple detectable oligonucleotides through reiterative enzymatic oligonucleotide synthesis events on a defined polynucleotide sequence. The methods generally comprise using a nucleoside, a mononucleotide, an oligonucleotide, or a polynucleotide, or analog thereof, to initiate synthesis of an oligonucleotide product that is substantially complementary to a target site on the defined polynucleotide sequence; optionally using nucleotides or nucleotide analogs as oligonucleotide chain elongators; using a chain terminator to terminate the polymerization reaction; and detecting multiple oligonucleotide products that have been synthesized by the polymerase. In one aspect, the invention provides a method for detecting a target protein, DNA or RNA by generating multiple detectable RNA oligoribonucleotides by abortive transcription.

The present invention relates to polyvinyl ethers and to their use as supports for synthetic methods (organic synthesis, peptide synthesis, oligonucleotide synthesis, oligosaccharide synthesis or any other synthetic procedure) and to their use for chromatographic applications (such as affinity chromatography), immobilisation of enzymes, reagents or catalysts.

The invention designs an untranslated region specific artificial micro RNA (miRNA) capable of effectively inhibiting replication of porcine reproductive and respiratory syndrome (PRRS) virus strains, which belongs to the field of researches on gene engineering and biological products. The sequence of the miRNA is represented by one of SEQ ID No.1, SEQ ID No.2, SEQ ID No.3 and SEQ ID No.4. The invention also discloses the construction of an artificial miRNA expression vector based on RNA polymerase II, design of miRRNAi, synthesis and cloning of double-stranded oligo, detection of expression of miRNA and inhibition effect on replication of different PRRSV strains. Compared with siRNAs for genes of other PRRSVs, the artificial miRNA screened by the invention, according to the conservative sequence of the untranslated region, can inhibit the replication of homologous virus strains and heterologous virus strains and can be used in both the development of anti-PRRSV infection preparations and breeding transgenic pig disease resistance lines.

Methods for encoding data in synthetic DNA include, for an input data sequence, constructing a codebook of a number of unique codewords, of a nucleotide length, which are constructed such that, for each codeword, the codeword is formed from a selection of nucleotide pairs from a first predefined dictionary of nucleotide pairs, and, if the nucleotide length of the codeword is odd, an additional selection of a nucleotide from a second predefined dictionary of individual nucleotides. For each symbol from the input data sequence, at least one codeword is designated as associable therewith, and each symbol is coded as one codeword selected from the designated at least one codeword associable with that symbol. A code is formed from the codewords, arranged in corresponding order to that of their respective symbols in the input data sequence. A DNA sequence is synthetized with nucleotides ordered to match the code.

The invention provides a method of convergently synthesizing mixtures of either single stranded or double stranded polynucleotides. In one aspect, oligonucleotides that form components of such polynucleotides are synthesized on one or more microarrays, or other large-scale parallel solid phase synthesis platforms, after which they are amplified directly, or are released into solution and then amplified. At least two sets of such released and amplified oligonucleotides are produced, referred to herein as first and second amplicons. The first and second amplicons are cleaved and then ligated to different ends of a bridging duplex that is present in the reaction in limiting quantity to form a polynucleotide mixture of the invention. At the completion of the reaction, each polynucleotide in the mixture is present in substantially equal concentration, regardless of the starting concentrations of the first and second amplicons. That is, the invention provides a method for synthesizing a normalized mixture of polynucleotides.

The invention relates to a recombinant bacterium for producing L-histidine, a construction method of the recombinant bacterium and a method of producing the L-histidine. Compared with an original strain, the recombinant bacterium for producing the L-histidine has the improved expression and / or activity of adenosine succinic acid synthetase PurA, and the original strain is a strain capable of accumulating the L-histidine. According to the recombinant bacterium and the construction method of the recombinant bacterium, based on the pioneering principle, by enhancing AMP synthesis in the synthesispath of nucleotide and improving the synthesis capacity of a bacterial strain ATP, enough precursor substances and energy carriers ATP are provided for efficient histidine synthesis, and the histidine synthesis capacity of the recombinant bacterium strain is significantly improved.

A polymer article having structure (I), where P comprises the basic polymer and groups -L-CH(-X(R1)p)-CH2(-Y(R2)p); L is a part of a pending group utilized for introducing -X(R1)p and -Y(R2)p; X and Y are halogen, N, S and O; R1 is hydrogen, alkyl, acyl or R2 when X is N, O or S; R2 is hydrogen, alkyl, alkylaryl or arylalkyl in which the alkyl part may contain 1-18 carbons, or -R3(-NH-CR4=O)q or -R3(-NH2)q, -CR4=O or poly alkyloxy that may have been terminally acylated or alkylated; p is an integer 0-3, with the provisos a) p depends on X being halogen, O, S or N, and b) that if several groups R2 are present they may be identical or different; R3 is alkyl, -O-alkyl, hydroxyalkyl, phenylalkyl, with up to 18 carbon atoms in the alkyl part, and R4 is C1-18 alkyl. q is an integer>O representing that one or more hydrogens in R3 may have been replaced with -NH2 or -NH-CR4=O. The polymer article may be used as a chromatography separation medium or in solide phase oligonucleotide synthesis.

The invention discloses an oligonucleotide synthesis chip system. The oligonucleotide synthesis chip system comprises an oligonucleotide synthesis chip and a synthetic carrier particle size screeningchip, wherein the high-throughput oligonucleotide synthesis chip comprises a micro-funnel structure with two ends open, an upper end opening is located in the upper surface of the high-throughput oligonucleotide synthesis chip, a lower end opening is located in the lower surface of the high-throughput oligonucleotide synthesis chip, and the caliber of the upper end opening is larger than that of the lower end opening. The synthetic carrier particle size screening chip comprises screen holes, wherein the upper surface opening diameter of the screen holes is smaller than the upper end opening aperture of the oligonucleotide synthesis chip, the lower surface opening diameter of the screen holes is obvious larger than the lower end opening diameter of the oligonucleotide synthesis chip, and the lower surface opening diameter of the screen holes is slightly smaller than the upper surface opening diameter of the screen holes.

The invention discloses a method for preparing an anti-EGFR safe chimeric antigen receptor modified immune cell and application thereof. The method comprises the following steps: synthesizing scFv(EGFR)-CD8-CD137-CD33 zeta-T2A-Leader-CD20-CD8-CD3 zeta-nucleotide; inserting a fusion gene fragment into a lentivirus expression vector, and packaging into a lentivirus carrying scFv(EGFR)-CD8-CD137-CD3zeta-T2A-Leader-CD20-CD8-CD3 zeta coding genes, and infecting a monocyte induced immune cell to obtain the anti-EGFR safe chimeric antigen receptor modified immune cell. According to the method, the safety of chimeric antigen receptor modified immune cell can be improved, toxic and side effect can be controlled, and occurrence of adverse response can be avoided.