48 results about "Ligase Gene" patented technology

The present invention relates to non-dependent ligase gene cloning process based on thio-modification. Each of the amplified target gene and the primer of the plamid vector has The 5' end thio-modified. The cloning process includes: PCR amplification of target gene and plamid vector and purification of PCR products; degrading DNA chain from the 5' end with 5' exonuclease and thio-modification to create 10-20 of 3' single chain raised ends of nucleoside; and direct transformation to colibacillus. The target gene cloning of the present invention has no dependence on restrictive endonuclease and DNA ligase and great operation flux, and may be used in large scale gene cloning.

The invention discloses a method of synthesizing L-carnosine using L-amino acid ligase by a one-step method. The method comprises the steps of taking beta-alanine, L-histidine, ATP (adenosine triphosphate) and polyphosphate as raw materials, and MgCl2 as an activator, adding L-amino acid ligase and polyphosphate kinase, and synthesizing L-carnosine through an enzymatic reaction and coupled coupleunder conditions of a pH of 6.5-8.5 and a temperature of 30-45 DEG C. According to the method, a sequence optimized L-amino acid ligase gene is successfully expressed in escherichia coli, and beta alanine and L-histidine can be catalyzed to synthesize L-carnosine at one step. A substrate is not required to be methylated or subjected to group protection; ATP required by a catalytic reaction is continuously regenerated by allowing polyphosphate kinase to catalyze phosphorylation of ADP (adenosine diphosphate); cyclic regeneration of ATP can be achieved only by consuming a small amount of ADP; and a renewable raw material, sodium hexametaphosphate, is wide in source, low in cost, simple to operate and easy in large-scale production.

A DNA fragment (8050 bp) for the 2-naphthalene acid degradating bacterium (Burkholderia sp. JT1500) and its preparing process are disclosed. Its prokaryotic expression plasmid is configured. Its recombinant bacterium strain containing said DNA fragment is obtained by transferring to cobacillus, which can effectively degradating 2-naphthalene acid and sodium benzoate, so degradating the environmental pollutants.

The invention provides a construction and fermentation method of an artificial strain with high yield of fengycin. The 4'-phosphoric acid pantetheinyl transferase gene sfp from bacillus amyloliquefaciens FZB42, a multi-effect factor gene degQ from bacillus subtilis 168 and a strong promoter P43 are integrated in series onto a genome of the bacillus subtilis 168 by utilizing CRISPR gene editing to construct a strain B.subtilis ds; an acetyl coenzyme A synthetase gene acs of Escherichia coli BL21, an acetyl coenzyme A carboxylase gene accACD of Salmonella enterica and a biotin ligase gene bisA of Corynebacterium glutamicum are connected in series to an expression vector pHT43, and the expression vector pHT43 is introduced into a fengycin synthesis strain B.subtilis ds, so that a fengycin high-yield strain B.subtilis dspabacd is constructed; the constructed engineering strain is fermented and cultured in a shake flask to produce the fengycin.

The present invention relates to a nitrogen-regulated RING-like ubiquitin E3 ligase gene required for sugar sensing and the modulation of the expression of this gene to modulate a characteristic in a plant. The RING-like ubiquitin E3 ligase of the present invention is involved in mediating nitrogen limitation adaptive responses in plants and its expression is influenced by nitrogen status. Increased expression of this or substantially similar genes can produce plants with improved nitrogen utilization and increased yield.

The present invention relates to a nitrogen-regulated RING-like ubiquitin E3 ligase gene required for sugar sensing and the modulation of the expression of this gene to modulate a characteristic in a plant. The RING-like ubiquitin E3 ligase of the present invention is involved in mediating nitrogen limitation adaptive responses in plants and its expression is influenced by nitrogen status. Increased expression of this or substantially similar genes can produce plants with improved nitrogen utilization and increased yield.

The invention relates to construction and fermentation optimization methods of an amino-bis-demethoxycurcumin high-yield strain. The invention discloses an escherichia coli engineering strain for synthesizing aminodidemethoxycurcumin by taking p-aminocinnamic acid as a substrate, and belongs to the field of synthetic biology or metabolic engineering. A synthetic route composed of a 4-coumaroyl coenzyme A ligase gene 4cl, a dimer ketone synthase gene dcs, a curcumin synthase gene curs3 and acetyl coenzyme A carboxylase genes accBC and dtsR1 is transferred into Escherichia coli MG1655 (DE3), andthe synthetic route is subjected to modular division and optimization to obtain an engineering bacterium with optimal yield. Through optimization of fermentation conditions, the yield of amino didemethoxycurcumin reaches 33mg / L after the strain is fermented for 48h by taking p-aminocinnamic acid as a substrate. The invention lays a foundation for industrial production and application of the high-activity amino didemethoxycurcumin.

The invention provided a method for preparing a bacterial polysaccharide-modified recombinant fusion protein and use of the bacterial polysaccharide-modified recombinant fusion protein. The method comprises: co-expressing a recombinant fusion protein and the Neisseria meningitidis O-oligosaccharyltransferase PglL in an O-antigen ligase gene-defective bacterium, and linking a polysaccharides endogenous or exogenous for the bacterium to the recombinant fusion protein by the O-oligosaccharyltransferase PglL, to obtain the bacterial polysaccharide-modified recombinant fusion protein. The protein can be used for preparing antibodies against bacterial polysaccharides and vaccines.

The invention relates to the technical field of biology, in particular to a tomato ubiquitination E3 ligase gene SlCHIP1, application and a gene SlCHIP2. The nucleotide sequences of the tomato ubiquitination E3 ligase genes SlCHIP1 and SlCHIP2 are cloned, the homology of the two genes, the evolutionary relationship with homologous genes of other species, and the effect and mechanism in high-temperature resistance of tomatoes are studied and discussed, and a new gene resource and a possible molecular marker are provided for high-temperature-resistant breeding and seed selection of the tomatoesand other species.

The invention discloses a gene engineering bacteria with high-yield malonyl coenzyme A and a construction method and application thereof. The gene engineering bacteria is constructed by: knocking outfive genes (ldhA, pta, frdA, poxB and adhE) in the Escherichia coli genome, and then introducing malonyl coenzyme A synthesis pathway genes including acetyl coenzyme A synthetase gene of the Escherichia coli, acetyl coenzyme A carboxylase gene of Salmonella enteritidis and biotin ligase gene of Corynebacterium glutamicum. According to the gene engineering bacteria of the invention, a highly-efficient accumulation of the malonyl coenzyme A can be realized by inhibiting internal acetyl coenzyme A outflow pathways of the engineering bacteria and constructing malonyl coenzyme A synthesis pathwaysinto different expression vectors to be transferred into the engineering bacteria; the engineering bacteria can efficiently synthesize a precursor, malonyl coenzyme A, of flavonoid compounds by takingacetic acid, a metabolite by-product of the Escherichia coli, as a substrate; and the engineering bacteria can be used to increase the yield of naringenin, a skeleton precursor of the flavonoid compounds, synthesized by a microbiological method.

The invention discloses a preparation method and application of beta-picric acid. The preparation method of beta-picric acid comprises a construction method of reconstituted cells, wherein the construction method of the reconstituted cells comprises steps (1), (2), (3) and (4) to reconstruct receptor cells so as to obtain the reconstituted cells: (1) introducing a picric acid synthesis related gene 1 into the receptor cells; (2) introducing a picric acid synthesis related gene 2 into the receptor cells; (3) introducing a short side chain ester-acyl coenzyme A ligase gene into the receptor cells; and (4) introducing a valerophenone synthetase gene into the receptor cells, wherein the picric acid synthesis related gene 1 is used for coding picric acid synthesis related protein 1, the picric acid synthesis related gene 2 is used for coding picric acid synthesis related protein 2, and the receptor cells are microbial cells, non-human animal cells or plant cells.

The invention discloses a dual-promoter universal plasmid for expressing T4 ligase of a domestic silkworm middle silk gland bioreactor as well as application and a method of the dual-promoter universal plasmid. The plasmid takes piggy Bac transposons as a foundation and carries an Amp resistance gene; the plasmid comprises function expression frames of a T4 ligase gene used as an exogenous gene and a green fluorescence EGFP (Enhanced Green Fluorescent Protein) gene used as a marker gene; the plasmid is constructed by utilizing a molecular biology method and two special restriction enzyme cutting sites containing ApaI and NheI are formed between DDDDK and a fibroin light-chain gene polyA; the universal plasmid is subjected to dual-enzyme digestion by adopting the ApaI and the NheI; after the universal plasmid is connected with the T4 ligase gene, the universal plasmid and an auxiliary plasmid are commonly injected into a domestic silkworm fertilized ovum; the green fluorescence protein gene and the T4 ligase gene are transferred into a domestic silkworm genome through utilizing properties of the transposons and are stably inherited and expressed to obtain transgenic domestic silkworms. The transgenic domestic silkworms are screened with the help of a fluorescence marker gene and domestic silkworm silk gland cells are used for specifically synthesizing and secreting T4 ligase protein.

The present invention relates to non-dependent ligase gene cloning process based on thio-modification. Each of the amplified target gene and the primer of the plamid vector has The 5' end thio-modified. The cloning process includes: PCR amplification of target gene and plamid vector and purification of PCR products; degrading DNA chain from the 5' end with 5' exonuclease and thio-modification to create 10-20 of 3' single chain raised ends of nucleoside; and direct transformation to colibacillus. The target gene cloning of the present invention has no dependence on restrictive endonuclease andDNA ligase and great operation flux, and may be used in large scale gene cloning.

A DNA fragment (8050 bp) for the 2-naphthalene acid degradating bacterium (Burkholderia sp. JT1500) and its preparing process are disclosed. Its prokaryotic expression plasmid is configured. Its recombinant bacterium strain containing said DNA fragment is obtained by transferring to cobacillus, which can effectively degradating 2-naphthalene acid and sodium benzoate, so degradating the environmental pollutants.

The invention discloses the application of soybean E3 ubiquitin ligase gene GmPUB2 for regulating plant flowering. Application of soybean E3 ubiquitin ligase gene GmPUB2 regulating plant flowering in regulating plant flowering. The GmPUB2 gene is transferred into the target plant through the plant expression vector to obtain the late flowering plant. The GmPUB2 gene is transferred into the target plant through the gene silencing vector to obtain the early flowering plant. The target gene GmPUB2 was transformed into Arabidopsis. Compared with wild-type Arabidopsis, it was proved that the bolting and flowering time of the transgenic plants were significantly later than those of the wild-type, indicating that the GmPUB2 gene has the effect of delaying plant flowering. Using the constructed GmPUB2 gene silencing vector, the transgenic Arabidopsis overexpressing the target gene GmPUB2 was silenced, and it was proved that the flowering time of the silenced plants was significantly earlier than that of the control plants.

The invention discloses a paulownia 4-coumaric-acid: coenzyme A ligase, encoding genes and application thereof. Amino acid residue sequence of the 4-coumaric-acid: coenzyme A ligase is shown in SEQ ID No. of 1. The information can be used for constructing genes of the 4-coumaric-acid: coenzyme A ligase, or antisense genes and RNA disturbing mass of the genes, which can be used for improving plantmaterials and constructing new materials with higher or lower lignin content.

The invention discloses a method of synthesizing L-carnosine using L-amino acid ligase by a one-step method. The method comprises the steps of taking beta-alanine, L-histidine, ATP (adenosine triphosphate) and polyphosphate as raw materials, and MgCl2 as an activator, adding L-amino acid ligase and polyphosphate kinase, and synthesizing L-carnosine through an enzymatic reaction and coupled coupleunder conditions of a pH of 6.5-8.5 and a temperature of 30-45 DEG C. According to the method, a sequence optimized L-amino acid ligase gene is successfully expressed in escherichia coli, and beta alanine and L-histidine can be catalyzed to synthesize L-carnosine at one step. A substrate is not required to be methylated or subjected to group protection; ATP required by a catalytic reaction is continuously regenerated by allowing polyphosphate kinase to catalyze phosphorylation of ADP (adenosine diphosphate); cyclic regeneration of ATP can be achieved only by consuming a small amount of ADP; and a renewable raw material, sodium hexametaphosphate, is wide in source, low in cost, simple to operate and easy in large-scale production.

The invention discloses a biological preparation method of typhoid glycoprotein and an application thereof. The invention discloses a method for preparing bacteria polysaccharide-modified protein, a substrate protein of neisseria meningitidis O-oligosaccharyltransferase PglL and the neisseria meningitidis O-oligosaccharyltransferase PgIL are co-expressed in a bacteria defective in O-antigen ligase genes, and the bacteria polysaccharide-modified protein is obtained. The biological preparation method has wide application prospect for increasing uniformity of vaccine, increasing the production efficiency of the vaccine, and reducing cost; and the vaccine can be used for preventing the diseases caused by typhoid pathogen.