86 results about "Polymerase Gene" patented technology

The invention relates to poly(ADP-ribose)polymerase (PARP) homologs which have an amino acid sequence which hasa) a functional NAD+ binding domain andb) no zinc finger sequence motif of the general formulaCX2CXmHX2C in which m is an integral value from 28 or 30, and the X radicals are, independently of one another, any amino acid;and the functional equivalents thereof; nucleic acids coding therefor; antibodies with specificity for the novel protein; pharmaceutical and gene therapy compositions which comprise products according to the invention; methods for the analytical determination of the proteins and nucleic acids according to the invention; methods for identifying effectors or binding partners of the proteins according to the invention; novel PARP effectors; and methods for determining the activity of such effectors.

The invention relates to the technical field of genetic engineering, and particularly relates to a method for preparing a gene recombinant expression enzyme of Thermusaquaticus TaqDNA polymerase. The nucleotide sequence of a gene and the amino acid sequence of an encoded protein are respectively shown as SEQIDNO:1 and SEQIDNO:2. According to the invention, the method comprises the following steps: cloning a gene sequence by using a gene cloning technique, and building a prokaryotic expression vector; by using a recombinant strain of a built TaqDNA polymerase gene, carrying out suspension by using an LB culture solution; after carrying out culturing for 4.5 hours at a temperature of 37 DEG C, adding IPTG (isopropyl-beta-d-thiogalactoside) with a final concentration of 0.5 mM into the obtained substance to induce for 16-20 hours; after thalli are collected in a centrifugal mode, dissociating the thalli by using 2-4 mg / ml lysozyme, so that protein fluid is obtained; and filtering the protein fluid by using a chromatographic column and a dialysis bag so as to obtain high-purity active protein TaqDNA polymerase. Compared with the prior art, stability of a production process is good, activity of obtained TaqDNA polymerase is high, the yield and purity of products are effectively ensured, and amplification effect when being applied to a gene PCR (polymerase chain reaction) is good.

The invention provides a genetic engineering bacterium for L-theanine production and a fermentation method thereof. A construction method of the genetic engineering bacterium comprises the steps of carrying out single copying on an ribonucleic acid (RNA) polymerase gene T7RNAP derived from T7 phage on an Escherichia coli W3110 genome, wherein the gene is controlled by a xylose promoter; carrying out dual copying on a gamma-glutamyl methylamine synthetase gene gmas derived from methylovorusmays, wherein the gene is controlled by a T7 promoter; knocking out a xylose operon repressor protein genexylR; and knocking out a succinyl CoA synthetase gene sucCD. The genetic engineering bacterium has the beneficial effects that the yield of L-theanine fermented by using the genetic engineering bacterium can be up to 40g / L and the sugar acid conversion rate can be up to 25%; and the L-theanine production method provided by the invention has the advantages of being low in raw material price, shortin cycle, simple and convenient to operate, green and friendly to environment and high in yield and has good industrial application value.

The invention discloses recombinant bacteria for producing salidroside and analogue thereof, and a construction method and use. The construction method comprises the steps of synthesizing keto-decarboxylase gene synkdc and glycosyl transferase gene synyjic, and connecting to a vector pRSFDuet-1 to construct a recombinant vector pSynkdc1-yjic; enabling T7RNA polymerase gene to be free or integrated to SyBE-002447 base bacterial strain, and introducing pSynkdc1-yjic to obtain SyBE-218011 and SyBE-218013 bacterial stains; and directly integrating synkdc and synyjic to SyBE-002447 chromosome, and removing feaB and ushA in a knockout manner to obtain a SyBE-218015 bacterial strain. The bacterial strain disclosed in the invention can be used for producing salidroside and analogue thereof through fermentation, so that the problem existing in source of the salidroside and analogue thereof can be solved, and cost can be lowered.

The invention discloses an iterative gene circuit based on a vibrio fischeri quorum sensing system and a T7 expression system. The iterative gene circuit comprises a signal molecular protein gene luxIof vibrio fischeri, a receptor protein gene luxR, a promoter sequence PluxI(1) for sensing a signal molecular and receptor protein complex, a T7 RNA polymerase gene T7 RNApoly and a green fluorescentprotein egfp for representing the circuit intensity. The gene circuit does not rely on an inducer and can utilize the T7 expression system to spontaneously and forcefully initiate target gene expression in any host bacterium; and gene elements are constructed on different expression vectors, the iterative gene circuit is constructed in engineering bacteria, the T7 expression system is utilized toamplify signals of the vibrio fischeri quorum sensing system, and the iterative gene circuit has the function of utilizing the T7 expression system to spontaneously and forcefully initiate target gene expression, and has a lower leakage ratio and higher initiating strength.

The invention discloses a construction method of a recombinant bacterium for producing short-and-medium-chain-length PHA and functional derivatives thereof. The method comprises the following steps of: introducing a coding gene of specific PHA polymerase, a coding gene of a key protein of a PHA synthesis pathway, a promoter or a mutated promoter, related genes capable of enhancing carbon source utilization capacity of medium-chain and long-chain fatty acids, the ribosome binding site RCJ and the ribosome binding site RD into a starting strain from which an endogenous PHA polymerase gene is knocked out; and adjusting the proportions of all monomers in the short-and-medium-chain-length PHA and the functional derivatives thereof in the recombinant bacteria so as to realize controllable production of the short-and-medium-chain-length PHA and the functional derivatives thereof.

The invention discloses a method and a kit for detecting genotypes of a herpes simplex virus (HSV). Primer probes designed on the basis of HSV DNA polymerase gene are adopted, type 1 and type 2 of the HSV are detected simultaneously in a single tube by TaqMan probe dual-wavelength fluorescent PCR technology, and the existence and the type of the HSV in a sample are judged through fluorescent signals and circulating threshold values of amplification templates with different wavelengths. In the kit, a manually constructed internal reference system is used for avoiding false negative of a detection result. The kit comprises nucleic acid extracting solution, PCR buffer solution, Taq enzyme, an internal reference, negative control and positive control; and two steps of sample treatment and amplification detection are adopted. The kit is easy and convenient to operate and high in sensitivity, can avoid false positive caused by PCR product nucleic acid pollution, can solve the problem of false negative caused by a PCR inhibitor in the sample, and can be widely applied to quick detection of the genotypes of the pathogen clinically.

The invention provides a strain capable of improving the performance of recombinant protein under an aerobic condition, and belongs to escherichia coli. The strain comprises at least one non-endogenous promoter which is operably connected to a glpF gene, a gldA gene, a dhaKLM gene operator sequence and an araE gene on the chromosome of the strain, an araBAD promoter which is operably connected to a T7RNA polymerase gene on the chromosome, and a T7 promoter which is operably connected to a recombinant protein gene. Furthermore, the strain is lack of an araABD gene operator sequence and an araFGH gene operator sequence.

The invention provides a genetic engineering bacterium for L-theanine production and a fermentation method thereof. A construction method of the genetic engineering bacterium comprises the steps of carrying out single copying on an ribonucleic acid (RNA) polymerase gene T7RNAP derived from T7 phage on an Escherichia coli W3110 genome, wherein the gene is controlled by a xylose promoter; carrying out dual copying on a gamma-glutamyl methylamine synthetase gene gmas derived from methylovorusmays, wherein the gene is controlled by a T7 promoter; knocking out a xylose operon repressor protein genexylR; and knocking out a succinyl CoA synthetase gene sucCD. The genetic engineering bacterium has the beneficial effects that the yield of L-theanine fermented by using the genetic engineering bacterium can be up to 40g / L and the sugar acid conversion rate can be up to 25%; and the L-theanine production method provided by the invention has the advantages of being low in raw material price, shortin cycle, simple and convenient to operate, green and friendly to environment and high in yield and has good industrial application value.