17 results about "Polymerase Gene" patented technology

The invention provides a genetically engineered bacterium for producing polydeoxyribonucleotide as well as a construction method and application of the genetically engineered bacterium. The genetically engineered bacterium comprises kluyveromyces marxianus for expressing a salmon DNA polymerase I gene. According to the invention, the edible probiotic Kluyveromyces marxianus is transformed through genetic engineering, efficient, safe and controllable production of PDRN is realized, the biological activity of PDRN is equivalent to that of PDRN from a natural source, and large-scale production and application of PDRN are facilitated.

The invention belongs to the technical field of genetic engineering, and discloses a plasmid-free genetically engineered bacterium for efficiently synthesizing gamma-aminobutyric acid from beginning by taking a cheap carbon source as a substrate, a method and application of the genetically engineered bacterium. The genetically engineered bacterium is obtained by overexpressing a T7RNA polymerase gene on the basis of a wild type E.coli MG1655; the gene gabT and the gene puuE are deleted; performing overexpression on the genes of gadC, gadbm, gdh, gltA, pyc and ppc; and the expression of sucA and argA genes in a GABA channel produced by escherichia coli is dynamically regulated and controlled by using a growth coupled promoter. The genetic manipulation of the engineering bacterium is carried out on a genome, plasmid residues, antibiotics and inducers are not used, the production performance is stable, and the fermentation process is simple. The engineering bacteria are used for fermenting for 38 hours under a staged pH value control process, the yield of gamma-aminobutyric acid can reach 35.4 g / L, and the engineering bacteria have a relatively good industrial application value.

The invention belongs to the field of genetic engineering and biological treatment, and provides an eukaryotic expression vector, gram-negative bacteria containing the eukaryotic expression vector and application of the eukaryotic expression vector in preparation of tumor drugs. By selecting and combining a phage-derived promoter, a specific phage RNA polymerase gene, a membrane rupture protein gene and an eukaryotic expression drug protein gene, an eukaryotic expression vector is divided into three modules: (1) a phage-derived RNA polymerase gene transcription module; (2) a membrane rupture protein gene expression module; and (3) a eukaryotic drug protein gene expression module. Through organic combination of the three modules, the eukaryotic drug delivery system is constructed, and the system and method for stably and efficiently performing eukaryotic drug protein expression, drug delivery and drug killing are realized.

The invention provides a preparation method of thermal-stability T7RNA polymerase and the thermal-stability T7RNA polymerase, belongs to the field of biology, and can solve the problems that an existing T7RNA polymerase preparation process is complex, the expression culture period is long, multi-step purification treatment is needed, liquid change treatment needs to be carried out independently before storage, and the production cost is high. The method comprises the following steps: transferring plasmids containing a T7RNA polymerase gene sequence into expression host bacteria M15, culturing, screening, cloning, adding glycerol for strain preservation, and establishing a strain three-level library; the method comprises the following steps: taking glycerol bacteria, carrying out streak resuscitation on a flat plate, selecting monoclone, inoculating the monoclone into an LB culture medium, culturing, carrying out enlarged culture, carrying out IPTG induced expression, centrifugally collecting thalli, cleaning, adding a lysis buffer solution, resuspending, carrying out ultrasonic cell disruption, centrifuging, and collecting supernate; purifying by nickel column affinity chromatography, removing impure protein by an impure washing buffer solution, eluting target protein, purifying by matching with anions and cations, and collecting the purified protein; and mixing the purified protein with a storage buffer solution, and subpackaging and storing. The method can be applied to rapid preparation of the thermal-stability T7RNA polymerase.

The present application relates to the field of genetic engineering and biological therapy, and provides a eukaryotic expression vector, a gram-negative bacterium containing the eukaryotic expression vector, and a use of the eukaryotic expression vector in the preparation of antitumor drugs and the treatment of tumors. By selecting and combining a phage-derived promoter, a specific phage RNA polymerase gene, a membrane-disrupting protein gene and a eukaryotic expression drug protein gene, the eukaryotic expression vector is divided into three modules: (1) a phage-derived RNA polymerase gene transcription module; (2) a membrane-disrupting protein gene expression module; and (3) a eukaryotic drug protein gene expression module. By means of organic combination of the three modules, a eukaryotic drug delivery system is constructed, and a system and method for stable and efficient eukaryotic drug protein expression, drug delivery and drug killing are implemented.

A kind of PLS DNA polymerase constructed by DNA polymerase-DNA binding protein and its application in DNA information storage belong to the technical field of biochemistry and molecular biology.The present application connects Sso7d DNA binding protein to the C-terminal of 9°N DNA polymerase by flexible short peptide to construct fusion protein PLS DNA polymerase.The codon-optimized PLS DNA polymerase gene is chemically synthesized, and high-purity PLS DNA polymerase is prepared by purification;the enzymatic properties of the PLS DNA polymerase and its application effects in in vitro DNA information storage addressing and in vivo DNA information storage assembling DNA active block are systematically evaluated.The results show that after fusing DNA binding protein, the performance of PLS DNA polymerase such as continuous synthesis ability, fidelity and PCR efficiency is significantly improved, and specific files can be recovered from mixed file pool to realize random addressing.

This invention belongs to the field of plant molecular biology, specifically relating to a small RNA molecule that alleviates lignification of loquat fruit and its application. The small RNA molecule is from loquat. Eja-MIR164 Its nucleotide sequence is shown in SEQ ID NO. 6. After processing within the plant, it forms a functional mature form. This invention experimentally confirms that loquat fruit contains… Eja-MIR164 The expression level of Eja-miR164 was negatively correlated with lignin content; transient interference or stable transformation using STTM technology reduced Eja-miR164 content, targeting its genes NAC transcription factor and lignin polymerase gene. EjLAC7 Increased expression levels in all samples led to a significant increase in lignin content. This invention utilizes small RNA molecules. Eja- MIR164 It can be used to breed new loquat varieties with reduced lignification, improve fruit texture and quality, and enhance post-harvest storage performance, and has potential application value in loquat genetic improvement and molecular breeding.

Disclosed are oligonucleotide primers that hybridize specifically with any base sequence designed from the base sequences of the N gene, RNA-dependent RNA polymerase gene, M gene, and S gene of SARS-COV-2, a nucleic acid amplification method using said primers, a test method for SARS-COV-2 infection by detection of nucleic acid amplification, and a COVID-19 test kit.

Provided are methods of treating a Primary Mitochondrial Disorder (PMD) or Mitochondrial DNA Depletion Syndrome (MDDS) in a subject in need thereof comprising: administering a therapeutically effective amount of a compound listed in Table 1, Table 2, Table 3, Table 4, or Table 5. The subject may have one or more mutations or deletions in the DNA polymerase γ gene (POLG).

Disclosed are oligonucleotide primers that hybridize specifically with any base sequence designed from the base sequences of the N gene, RNA-dependent RNA polymerase gene, M gene, and S gene of SARS-CoV-2, a nucleic acid amplification method using said primers, a test method for SARS-CoV-2 infection by detection of nucleic acid amplification, and a COVID-19 test kit.

Provided are methods of treating a subject having a Primary Mitochondrial Disorder (PMD) or Mitochondrial DNA Depletion Syndrome (MDDS) comprising administering a therapeutically effective amount of a compound listed in Table 1, Table 2, Table 3, Table 4, or Table 5, wherein the subject does not have a mutation in the DNA polymerase γ gene (POLG).

The invention discloses a fluorescent quantitative PCR (Polymerase Chain Reaction) primer group and a detection kit for detecting type-2 agama herpesvirus and application of the fluorescent quantitative PCR primer group and the detection kit. According to the invention, a pair of specific primers IghV2-F and IghV2-R are designed for a gene conserved region of DNA-dependent DNA polymerase (pol) of iamyl herpesvirus type 2 (IghV2), and an SYBR Green dye fluorescent quantitative PCR detection method is established, so that the primer has the advantages of high sensitivity, high specificity, wide linear range, rapidness, simplicity, convenience, good repeatability and the like. The invention provides important technical support for herpesvirus infection monitoring, quarantine and protection work of rare and endangered lizard species, fills the blank of detection methods in the field, and is suitable for non-clinical medical fields such as scientific research, epidemiological investigation, quarantine monitoring, protection management and the like.

The invention discloses an RNA polymerase intracellular mutagenesis system and a construction method thereof. The mutagenesis system comprises a mutagenesis plasmid, a target gene sequence plasmid (GOI plasmid) and a reporter plasmid, the mutagenic plasmid carries an inducible promoter, a DNA polymerase gene sequence, an error-prone DNA polymerase gene sequence and an auxiliary protein gene sequence thereof; the GOI plasmid carries a constitutive strong promoter and a T7 RNA (Ribonucleic Acid) polymerase; the report system carries an inducible promoter and a resistance gene sequence. The plasmid system can perform continuous mutagenesis on a target sequence, the mutagenesis condition is visualized through report plasmids, the whole mutagenesis screening process has the characteristics of high throughput, low cost, simplicity in operation and the like, flora capable of growing in a high-concentration resistant environment can be obtained through simple passage screening, and the plasmid system has a wide application prospect. The construction difficulty of a traditional mutant library is greatly reduced.

A constitutive expression vector containing dual bacteriophage-derived promoters, a gram-negative bacterium containing the expression vector, and uses thereof in preparation of tumor drugs and treatment of tumors. By selecting bacteriophage-derived promoters, a specific bacteriophage RNA polymerase gene, a membrane-disrupting protein gene, and a therapeutic protein gene, the expression vector is divided into a protein expression control module, a drug delivery module, and a drug-mediated killing module. By organically combining the three modules, a three-in-one system is formed, and a system and method for stable and efficient drug protein expression, drug delivery, and drug-mediated killing are realized.

Methods are disclosed for high throughput identification of optimized template-independent polymerase variants. The methods comprise generation of polymerase gene-encoding variants, isolation of variants, expression of variants, exposing expressed polymerase variants to desired conditions for activity, and identification of active variants, such as by hybridization to synthesized polynucleotides or by amplification. Encoding genes of active variants can be sequenced. The method can be performed iteratively to enhance generation and selection of preferred variants.

Solid supports and methods using the solid supports for de novo long oligo synthesis are disclosed. Long oligos with more than 100 nucleotides are notably difficult to synthesize. The challenges include low percentage yield, high error rate, and difficulty to isolate the correct sequence. Using the disclosed solid supports and their associated methods, oligos with more than 1,000 nucleotides can be readily obtained. The synthesis of the 1,728-nucleotide Φ29 DNA polymerase gene was demonstrated.

The invention discloses a construction method of recombinant escherichia coli for efficiently synthesizing pterostilbene, and belongs to the technical field of biology. The method comprises the following steps: firstly, inserting an RNAP polymerase gene segment into a safe site SS9 of an escherichia coli genome, then integrating a dCas9 gene into a ycdn site of a recombinant bacterium, then transferring into a FabF-sgRNA plasmid or / and a fumC-sgRNA plasmid, dynamically regulating the malonyl-CoA level, and constructing a pathway for synthesizing pterostilbene from L-tyrosine, thereby obtaining the pterostilbene. By using a co-localization strategy of the enzyme, the accumulation of intermediate metabolites is further reduced, the catalytic efficiency of the enzyme is improved, the yield of the escherichia coli for synthesizing the pterostilbene from the glycerol is improved, the concentration of other byproducts is obviously reduced, and the method has a very good application prospect.