107 results about "RNA polymerase" patented technology

The invention provides an RNA polymerase variant and application thereof, and relates to the technical field of biology. Wild type T7 RNA polymerase is modified to obtain the T7 RNA polymerase variant, the enzyme specific activity of the T7 RNA polymerase variant is remarkably improved compared with that of the wild type T7 RNA polymerase, the addition amount of the polymerase in the in-vitro transcription reaction process can be reduced, and the cost is saved; in addition, the mutant also effectively reduces the generation of incomplete fragment impurities, and the mRNA integrity is significantly improved to 99%. The invention further provides a method for preparing RNA through in-vitro transcription.

The invention discloses an oil palm U6 promoter gene and application thereof, and belongs to the technical field of biology. The nucleotide sequence of the promoter gene is shown as SEQ ID NO.1. The oil palm RNA polymerase III type promoter gene, namely the oil palm endogenous U6 promoter gene EgU6, is obtained by cloning in an oil palm genome for the first time, and the promoter gene has high transcriptional activity and can drive downstream fluorescent protein mNeonGreen expression. The candidate oil palm endogenous U6 promoter gene can be provided for subsequently establishing a high-efficiency oil palm gene editing technology system based on a CRISPR / cas9 (Clustered Regularly Interspaced Short Palindromic Repeats / Cas9) system.

The invention provides an RNA polymerase variant, the enzyme specific activity of the variant is obviously improved compared with that of a wild type, and the addition amount of polymerase is reduced, so that the cost is saved. In addition, the invention further provides a method for generating RNA through in-vitro transcription.

The invention provides an RNA polymerase variant and application thereof, and relates to the technical field of biology. The RNA polymerase variant provided by the invention has the performance of remarkably improving the integrity of a super-long fragment mRNA transcription product, can be used for preparing mRNA through in-vitro transcription, particularly can improve the integrity of the product when preparing saRNA exceeding 10000 nt, and has important significance for industrial production of saRNA.

The invention provides a kind of RNA polymerase variants and application thereof, the variants have improved catalytic activity compared with wild type T7RNA polymerase, and the capping rate of mRNA products in the in vitro transcription (co-transcription capping) process can be increased. In addition, the disclosure also provides a method for preparing RNA by using the variant, and by using the method to prepare RNA molecules, more capped mRNA can be obtained.

The invention provides an RNA (Ribonucleic Acid) polymerase variant and a preparation method thereof, and an RNA product with low dsRNA pollution can be obtained by applying the RNA polymerase variant disclosed by the invention to in-vitro transcription. In addition, the invention further provides a method for preparing RNA through in-vitro transcription.

This invention generally relates to a novel RNA / mRNA production and amplification method using viral RNA replicase and / or RNA-dependent RNA polymerase (RdRp) enzymes as well as the associated mRNAs thereof. The present invention can be used for manufacturing and amplifying all varieties of RNA / mRNA sequences carrying at least an RdRp-binding site in the 5′- or 3′-end, or both. The RNA / mRNA so obtained is useful for not only producing mRNA vaccines and / or RNA-based medicines but also for generating the mRNA-associated proteins, peptides, and / or antibodies under an in-vitro as well as in-cell translation condition. Principally, the present invention is a novel RNA replicase-mediated RNA / mRNA amplification method, namely Replicase Cycling Reaction (RCR). The RNA replicases involved in RCR include but not limited to viral and / or bacteriophage RNA-dependent RNA polymerases (RdRp), particularly coronaviral and hepatitis C viral (HCV) RdRp enzymes.

The invention belongs to the technical field of biology, and relates to an RNA polymerase variant, and when the RNA polymerase variant is applied to in-vitro transcription, an RNA product with low dsRNA pollution can be obtained. In addition, the invention further provides a method for generating RNA through in-vitro transcription.

The present disclosure provides variant RNA polymerases with amino acid substitutions, and their use of which increases transcription yield, RNA integrity, capping efficiency, and reduces dsRNA formation during an in vitro transcription reaction.

The invention provides the RNA polymerase variant and the preparation method thereof, the RNA polymerase variant is applied to in-vitro transcription, an RNA product with low dsRNA pollution and high integrity can be obtained, meanwhile, the utilization efficiency of cap analogues in a co-transcription capping reaction system can be improved, and the cost is saved. In addition, the invention further provides a method for generating RNA through in-vitro transcription.

The invention provides an RNA polymerase variant and application thereof, and relates to the technical field of biology. According to the present invention, the wild type T7 RNA polymerase is modified to obtain the T7 RNA polymerase variant, and the capping rate of the RNA product can be substantially improved compared with the wild type T7 RNA polymerase;

This application provides an RNA polymerase variant and its application in in vitro transcription to prepare RNA, relating to the field of biotechnology. Using the RNA polymerase variant provided in this application for in vitro transcription not only reduces the generation of truncated or over-extended impurities in low-magnesium in vitro transcription reaction systems, thus improving the integrity of the transcribed RNA product, but also increases the yield and capping rate of the RNA product in low-magnesium in vitro transcription reaction systems. Furthermore, this application also provides the application of the RNA polymerase variant in in vitro transcription.

The methods, compositions, and kits of the disclosure provide a novel approach for a whole genome, unbiased DNA analysis method that can be performed on limited amounts of DNA can be used to analyze DNA to determine its modification status. Aspects of the disclosure relate to a method for amplifying bisulfite-treated deoxyribonucleic acid (DNA) molecules comprising: (a) ligating an adaptor to the DNA molecules, wherein the adaptor comprises a RNA polymerase promoter comprising bisulfite-protected cytosines; (b) treating the ligated DNA molecules with bisulfite; (c) hybridizing the bisulfite-treated DNA molecules with a primer; (d) extending the hybridized primer to make double stranded DNA; and (e) in vitro transcribing the double-stranded DNA to make RNA.

Provided is an RNA polymerase mutant in which an amino acid residue selected from the group consisting of glutamic acid at position 48 and arginine at position 50 in the amino acid sequence of a wild-type T7 RNA polymerase set forth in SEQ ID NO: 1 is substituted with an aliphatic amino acid residue.

The invention relates to a Pseudomonas sp. strain for use in the production of a recombinant protein characterised in that said strain comprises a nucleotide sequence encoding a phi15 RNA polymerase. The invention further relates to a plasmid, capable of integrating or replicating in Pseudomonas sp., comprising a phi15 promoter sequence operably linked to a nucleotide comprising one or more restriction sites for the insertion of a nucleotide sequence encoding a recombinant protein, or operably linked to a nucleotide sequence encoding a recombinant protein.

This invention belongs to the field of biology, specifically relating to a composition and its use against foot-and-mouth disease virus (FMDV). This invention discovered that compounds Pasireotide and Oxyclozanide alone cannot significantly inhibit FMDV replication. Unexpectedly, the combined use of Pasireotide and Oxyclozanide significantly inhibits the expression of FMDV RNA polymerase 3D protein, reducing the half-maximal tissue-cell infection dose (TCID) of FMDV. 50 It can significantly reduce the replication ability of foot-and-mouth disease virus and has the effect of antagonizing foot-and-mouth disease virus infection; and there are currently no clear clinical studies or drug instructions mentioning that there is a direct physical / chemical incompatibility between the two, so it can be used to prepare drugs against foot-and-mouth disease virus and has good application prospects.

The invention provides the RNA polymerase variant and the preparation method thereof, the RNA polymerase variant is applied to in-vitro transcription, an RNA product with low dsRNA pollution and high integrity can be obtained, meanwhile, the utilization efficiency of cap analogues in a co-transcription capping reaction system can be improved, and the cost is saved. In addition, the invention further provides a method for generating RNA through in-vitro transcription.

This invention discloses a method and kit for detecting RNA using a three-stage amplification melting curve based on isothermal self-circulation. The method includes designing a first primer, a second primer, and a circular amplification probe targeting the target nucleic acid sequence. The first primer consists of an RNA polymerase promoter sequence and a target nucleic acid-specific binding sequence, sequentially from the 5' end to the 3' end. The circular amplification probe is a circular DNA segment whose sequence consists of three parts: the first part completely contains a continuous sequence composed of the tag sequence and the nicking enzyme-specific recognition sequence from the second primer; the second part is a self-cleaving DNase sequence. In the technical solution used in this invention, a cascaded nucleic acid amplification system is constructed, achieving exponential signal amplification of the target nucleic acid through the synergistic effect of rolling circle amplification and RNA polymerase-mediated transcriptional amplification.

The invention provides the RNA polymerase variant and the preparation method thereof, the RNA polymerase variant is applied to in-vitro transcription, an RNA product with low dsRNA pollution and high integrity can be obtained, meanwhile, the utilization efficiency of cap analogues in a co-transcription capping reaction system can be improved, and the cost is saved. In addition, the invention further provides a method for generating RNA through in-vitro transcription.

The utility model relates to the technical fields of kit, put forward a kind of mRNA in vitro transcription kit, kit inside is separated into storage cavity and refrigeration cavity by partition, the refrigeration cavity inside is equipped with refrigeration device, the storage cavity inside includes the support seat fixed in the upper portion of partition, rotatable connection has the rotating shaft of vertical arrangement on the support seat, the top of rotating shaft coaxially is fixed with the rack for placing reagent tube for placing, driving hole is opened along the axis in the rack, the driving hole passes through rotating shaft and extends to the inside of support seat, spiral rod is passed in the inside of driving hole, the driving hole is located on the inner wall of rack and is opened with the spiral groove of the appearance matching of spiral rod. The utility model has the beneficial effects that: using spiral rod and refrigeration device, RNA polymerase can be centrifuged in kit, wait for use, do not need to take reagent out of box again, reduce the risk of temperature rise enzyme activity decline.

The invention provides the RNA polymerase variant and the preparation method thereof, the RNA polymerase variant is applied to in-vitro transcription, an RNA product with low dsRNA pollution and high integrity can be obtained, meanwhile, the utilization efficiency of cap analogues in a co-transcription capping reaction system can be improved, and the cost is saved. In addition, the invention further provides a method for generating RNA through in-vitro transcription.

This invention provides an RNA polymerase variant that can significantly reduce the generation of dsRNA impurities during in vitro transcription. The invention also relates to compositions and kits comprising this variant, and the use of the variants, compositions, and kits described herein in in vitro transcription.

This invention discloses an antisense oligonucleotide targeting and inhibiting the RdRp gene and its applications. This study designed and screened a series of antisense oligonucleotides (ASOs) targeting the highly conserved SARS-CoV-2 RNA-dependent RNA polymerase (RdRp), and systematically evaluated their antiviral effects against various SARS-CoV-2 variants. Experimental results show that the designed ASO molecules, especially RDRP-5, exhibit significant viral inhibitory activity at multiple experimental levels (including reporter gene systems, viral replicon models, and true viral infection experiments).

The application provides a T7 RNA polymerase variant and application thereof, wherein the amino acid sequence of the T7 RNA polymerase variant comprises at least one substitution or deletion of an amino acid site selected from R34, K172, Y178, R386, D388, Q435, N437, D438, relative to SEQ ID NO: 1. The variant has improved catalytic activity relative to wild-type T7 RNA polymerase, and can improve the capping rate of mRNA products in the process of in vitro transcription (cotranscriptional capping). In addition, the application also provides a method for preparing RNA by using the variant, and more capped mRNA can be obtained by using the method for preparing RNA molecules.

The present disclosure relates to engineered RNA polymerase polypeptides and compositions thereof, and polynucleotides encoding the engineered RNA polymerase polypeptides. The present disclosure also provides methods of producing RNA using the engineered RNA polymerase polypeptides or compositions thereof.

The invention belongs to the field of biology, and particularly relates to a composition for resisting foot and mouth disease viruses and application. The invention finds that single compounds of Pasireotide and Oxyzane cannot significantly inhibit the replication of the foot-and-mouth disease virus, unexpectedly, the combined use of the compounds of Pasireotide and Oxyzane can significantly inhibit the expression of the 3D protein of the RNA polymerase of the foot-and-mouth disease virus, reduce the half histiocyte infection dose (TCID50) of the foot-and-mouth disease virus, significantly reduce the replication ability of the foot-and-mouth disease virus, and improve the therapeutic effect of the foot-and-mouth disease virus. The effect of antagonizing foot-and-mouth disease virus infection is achieved; at present, no clear clinical research or drug specification means that the two compounds have direct physical / chemical incompatibility, can be used for preparing the anti-foot-and-mouth disease virus drugs, and have a good application prospect.

This invention provides an improved in vitro transcription method that produces early-termination transcripts and full-length mRNA transcripts that do not contain double-stranded mRNA. [Solution] The present invention provides a method for preparing DNA sequences optimized as templates for in vitro transcription of mRNA. These DNA sequences are optimized to avoid premature termination of transcription by RNA polymerase. The present invention also provides a method for preparing optimized DNA sequences containing one or more termination signals at the 3' end to reduce or prevent non-template "run-off" transcription.

The present disclosure relates to engineered RNA polymerase polypeptides and compositions thereof, and polynucleotides encoding the engineered RNA polymerase polypeptides. The present disclosure also provides methods of producing RNA using the engineered RNA polymerase polypeptides or compositions thereof.

The present disclosure provides methods and compositions for improving the in vitro transcription (IVT) of messenger RNA (mRNA) using SP6, T7, or KP34 RNA polymerase. In particular, the present disclosure provides in vitro transcription templates with optimized nucleic acid sequences specific for use with SP6, T7, or KP34 RNA polymerase that improve mRNA transcript yield and / or reduce amounts of dsRNA.