159 results about "Microbial Genes" patented technology

The invention relates to a saccharomyces cerevisiae strain for efficiently synthesizing cordycepin as well as a construction method and application of the saccharomyces cerevisiae strain, and belongs to the technical field of microbial genetic engineering. The construction method provided by the invention comprises the following steps: carrying out multi-copy genome integrated expression on heterologous codon optimized 2 '-carbonyl-3'-deoxyadenosine reductase genes and 3 '-adenosine monophosphate phosphohydrolase genes, so as to increase the dosage of key enzymes; and supplementing the defective gene of the engineering strain to obtain the saccharomyces cerevisiae strain B5U3 with high yield of cordycepin. According to the obtained saccharomyces cerevisiae strain for efficiently synthesizing the cordycepin, the cordycepin yield reaches 2.92 g / L after shake flask fermentation, the cordycepin yield reaches 7.5 g / L after fermentation in a fermentation tank, and a foundation is laid for safer and more reliable industrial production of the cordycepin.

The invention relates to the technical field of microbial genetic engineering, in particular to a method for improving cordycepin synthesized by saccharomyces cerevisiae and saccharomyces cerevisiae engineering bacteria. According to the invention, codon-optimized 2 '-carbonyl-3'-deoxyadenosine reductase gene and 3 '-adenosine monophosphate phosphohydrolase gene from cordyceps militaris are expressed by using plasmids, and then the plasmids are transformed into host cells to obtain the saccharomyces cerevisiae engineering bacteria. And inoculating a seed solution obtained by seed culture of the activated saccharomyces cerevisiae engineering bacteria into a fermentation culture medium containing a metabolic effector, and carrying out fermentation culture to synthesize cordycepin. Metabolic effectors (Cu < 2 + >, Fe < 2 + >, Mg < 2 + >, Zn < 2 + >, citric acid, cysteine, aspartic acid, glycine, VB1 and tea polyphenol) are added into a fermentation culture medium strain, and the variety and concentration of the metabolic effectors are optimized, so that the yield and synthesis efficiency of cordycepin are remarkably improved. By using the combination of the strain obtained by the invention and the optimal effector, the yield of cordycepin in a fermentation tank reaches 2.9 g / L.

The present invention discloses a digital analysis method based on off-site monitoring of water pollution source tracing, comprising the following steps: S1, collecting water samples: collecting water samples at multiple predetermined sampling points in the target water area, S2, environmental DNA extraction: performing environmental DNA extraction operation on the collected water samples, S3, microbial gene detection: performing microbial gene detection on the extracted environmental DNA using high-throughput sequencing technology, S4, data analysis and model construction: performing data analysis based on multiple algorithms, S5, pollution source tracing determination: combining satellite remote sensing data, the water color, area changes, algae distribution, floating objects on the water surface and other information of the water area can be monitored.

The invention belongs to the technical field of microbial genetic engineering, and particularly relates to an alpha-2, 3-sialyltransferase mutant and application thereof. The a-2, 3-sialyltransferase mutants V175N, P198A, G173D, D174Q, F196K and A315H are obtained by performing site-specific mutagenesis on six amino acids in an a-2, 3-sialyltransferase amino acid sequence coded by a gene derived from Campylobacter jejuni (OH4384), the yield of 3 '-sialyllactose is remarkably increased by single-point mutagenesis of the V175N, the yield of 3'-sialyllactose is remarkably increased by single-point mutagenesis of the V175N, the yield of 3 '-sialyllactose is remarkably increased by single-point mutagenesis of the P198A, the G173D, the D174Q, the F196K and the A315H, the optimal recombinant microbial strain 3 '-SL-09 (V175N / P198A / D174Q) is obtained through combination of multiple mutation sites, the yield can reach 5.91 g / L, the bottleneck problem of low catalytic efficiency of sialyltransferase is effectively solved, and an enzyme element with excellent performance is provided for high-efficiency and low-cost biological manufacturing of 3'-SL.

The invention belongs to the field of microbial genetic engineering, and provides schizochytrium limacinum with high DHA content and low DPA content and application of the schizochytrium limacinum. According to the strain, an n3 front-end desaturase gene derived from crypthecodinium cohnii is integrated to a 18S rDNA site of schizochytrium limacinum ALS01, and omega-6 type DPA is converted into DHA by utilizing an extension-desaturation way. Under the optimized conditions of a 5L fermentation tank (the inoculation amount is 5-20%, the temperature is 25-28 DEG C, the ventilation ratio is 1.0-2.0 vvm, and fermentation is performed for 72-168 hours), the biomass is larger than or equal to 150g / L, the total grease is larger than or equal to 60%, the DHA total grease proportion is larger than or equal to 53%, the DPA total grease proportion is smaller than or equal to 2%, and the DHA yield is larger than or equal to 47.82 g / L. The invention solves the problems of unstable fish oil resource and insufficient DHA purity in the traditional DHA production, provides a new strain for the industrial production of high-purity DHA, and has important application value.

The invention discloses a methane leakage microbial marker mining method based on multi-source data fusion and related equipment, according to the embodiment of the invention, a multi-dimensional feature space is constructed by fusing microbial gene data, spatial information and environmental geochemical data, and the data dimension and reliability of marker mining are improved; moreover, according to the embodiment of the invention, a mode of combining random forest and regression modeling is adopted, so that quantitative screening of the markers is realized, and manual intervention and subjective deviation are reduced; specifically, according to the embodiment of the invention, the markers are graded on the basis of comprehensive scores of three dimensions of importance, stability and universality, so that priority selection in subsequent practical application is facilitated; the embodiment of the invention is suitable for methane leakage identification of different sea areas and different geological backgrounds, has good generalizability and practicability, and can be widely applied to the technical field of marker analysis.

The invention discloses a pathogenicity-related sclerotinia sclerotiorum gene SsVSP19 and application thereof, and belongs to the technical field of microbial genetic engineering, the pathogenicity-controlling gene SsVSP19 from sclerotinia sclerotiorum provided by the invention has a DNA sequence as shown in SEQ ID No: 1, and consists of 1298 nucleotides; the amino acid sequence of the protein coded by the provided SsVSP19 gene is as shown in SEQ ID No: 2, and the protein consists of 332 amino acids; the SsVSP19 gene can be applied to the field of plant sclerotinia sclerotiorum gene engineering. The gene SsVSP19 for controlling pathogenicity of sclerotinia sclerotiorum is knocked out, so that the pathogenicity of the gene SsVSP19 has defects, and the gene SsVSP19 can be applied to reducing the pathogenicity of sclerotinia sclerotiorum.

Agriculturally useful genetically engineered bacteria which express RNAs or proteins that produce agriculturally useful compounds including ammonia, phosphate, and pesticides upon decreases in phosphate concentrations in plant growth media and methods of their use are disclosed.

The invention belongs to the technical field of microbial genetic engineering, and particularly relates to an alpha-2, 6-sialyltransferase mutant and application thereof. The a-2, 6-sialyltransferase mutants M133L, E140D, D228H, N289D and G322Q are obtained by carrying out site-specific mutagenesis on five amino acids in an a-2, 6-sialyltransferase amino acid sequence coded by a gene derived from Photobacterium sp.JT-ISH-224, and compared with wild type alpha-2, 6-sialyltransferase, the yield of the single-point mutant 6 '-SL of the E140D and the yield of the single-point mutant 6'-SL of the G322Q are increased by 87.80% and 78.01% respectively. The yield of the optimal recombinant microbial strain 6 '-SL-7 (E140D / G322Q) obtained by mutation site combination can reach 1.471 g / L, the bottleneck problem of low catalytic efficiency of sialyltransferase is effectively solved, and an enzyme element with excellent performance is provided for high-efficiency and low-cost biological manufacturing of 6'-SL.

The invention relates to a construction method and application of recombinant escherichia coli for increasing the yield of lactoyl-N-tetrasaccharide, and belongs to the field of microbial genetic engineering. According to the invention, three genes tauB, nikD and livM screened by using a transcriptome technology are proved to be capable of promoting the generation of LNT and LNTri II. An engineering strain MG-LNT-09 is used as a starting strain, the biosynthesis efficiency of endogenous transporter genes tauB, nikD, livM, LNT and LNTri II derived from Escherichia coli K-12 derivative bacteria MG1655 is remarkably improved through single plasmid overexpression, the yield of the LNT in shake-flask culture is as high as 7.77 g / L, and the yield of the LNTri II is as high as 3.03 g / L. The optimized strain MG3WB of which the tauB gene is integrated to the hlyE site obtains the maximum LNT titer of 40.35 g / L in a 5L bioreactor without using any antibiotics or plasmids, has a good industrial prospect, and lays a foundation for further industrial production and synthesis of other complex breast milk oligosaccharides.

The invention discloses a method for producing phloretin by using saccharomyces cerevisiae, and belongs to the technical field of microbial genetic engineering. A natural metabolic phloretin path in a plant is successfully constructed in the saccharomyces cerevisiae BY4741, synthesis of phloretin in microorganisms is achieved, synthesis of the phloretin in the microorganisms is enhanced by enhancing synthesis path genes of tyrosine and phenylalanine in the saccharomyces cerevisiae, including elimination of tyrosine feedback inhibition and introduction of exogenous genes, synthesis of prerequisite tyrosine and phenylalanine is enhanced, and the yield of the phloretin is increased. And the yield of the phloretin is further improved by enhancing the expression of a key gene in a phloretin synthesis pathway. The yield of phloretin synthesized from the beginning by the engineering yeast can reach 246.21 mg / L, the synthesis problem of phloretin serving as a carbohydrase inhibitor is effectively solved, and a foundation is laid for synthesizing phloretin through efficient fermentation of microorganisms.

The invention discloses a Stemphylium alcohol dehydrogenase as well as a coding gene and application thereof, and relates to the technical field of microbial genetic engineering. The alcohol dehydrogenase is any one of (1), (2) and (3): (1) SvADH1 protein, the amino acid sequence of which is as shown in SEQ ID NO.3; (2) carrying out substitution, deletion and / or addition of one or more amino acid residues on the amino acid sequence of the SvADH1 protein to obtain a protein which has more than 70% of identity with the SvADH1 protein and is related to cross-linked solanol A redox; and (3) a fusion protein obtained by connecting a protein tag to the N terminal or / and C terminal of (1) or (2). Research finds that overexpression of the coding gene of the alcohol dehydrogenase in Stemphylium cystosum can significantly improve the yield of the interlinked solanol A, so that a new direction is provided for developing the interlinked solanol A from microorganisms.

A microbial gene editing scheme automatic design method, system and storage medium; the method comprises: selecting a design object for different types of strains, the design object is bacteria or fungi; after determining the target gene, obtaining the target gene information from the NCBI database; the design scheme of bacteria comprises the following steps: determining the gRNA design region; selecting the optimal gRNA sequence; doing sequence complexity and GC content analysis; the design scheme of fungi uses a frameshift scheme: the gRNA of the frameshift scheme is set at any position not overlapping with other genes after the start codon in the gene; select gRNA sequence in gRNA design region, select the optimal gRNA sequence; design two pairs of four primer sequences for homologous arm identification primer; this scheme can replace the complicated and time-consuming manual scheme, save a lot of time, solve the problem of high cost in the design of the best site and the design of sgRNA sequence scheme of artificial gene knockout.

This invention relates to the fields of microbial genetic engineering and fermentation biomanufacturing technology, specifically disclosing a recombinant Corynebacterium glutamicum with high 5-aminolevulinic acid (ALA) production and its applications. Addressing the problems of existing strains such as crude enzyme activity regulation, inhibited cell growth, the need for external inhibitors, and low yield, this invention uses Corynebacterium glutamicum as a host and removes the gdhA gene without scarring to reduce precursor competitive metabolism. Site-directed mutations of the endogenous hemB gene (S288A, D128E, R231K) are performed to moderately reduce dehydratase activity. A high-expression plasmid pXMJ19-hemA containing the hemA gene from Rhodopsinus capsulatum is constructed and introduced into the modified strain to obtain a recombinant strain with optimized triple metabolism. This strain reduces product degradation without the need for external dehydratase inhibitors, does not affect cell growth, and achieves a yield up to 4.30 times higher than the control after 42 hours of fermentation, with ALA accumulation exceeding 8.9 g / L. The strain exhibits stable characteristics, the process is simple, and it has good industrial application value.

The invention belongs to the field of microbial genetic engineering. The invention particularly relates to a tylosin industrial high-yield strain as well as a construction method and application thereof. The sequence of the ALDH interfering gene is as shown in SEQ ID NO: 3. The industrial high-yield strain of tylosin is obtained by regulating and controlling the ALDH gene of an original strain. The strain is applied to fermentation production of tylosin and related derivatives thereof. The CRISPR i technology is utilized to inhibit in streptomyces fradiae to regulate and control ALDH gene expression, the gene is knocked out by using the homologous recombination technology, and then an engineering strain for high-yield tylosin is obtained. The engineering strain can improve the fermentation yield of tylosin by 6.8% and 9.9%.

The present invention belongs to the technical field of microbial genetic engineering, and particularly relates to an engineered strain for producing 2'-deoxyadenosine, a preparation method thereof and an application thereof. The engineered strain expresses a ketodeoxyadenosine reductase gene cns1 and an adenylic acid dephosphatase gene cns2 . The engineered strain prepared by the preparation method of the present invention has obvious advantages in the production of 2'-deoxyadenosine. By optimizing the fermentation medium conditions, the present invention improves the yield and efficiency of Corynebacterium glutamicum in synthesizing 2'-deoxyadenosine to varying degrees, and the yield reaches 702.3 mg / L on a 6L bioreactor.

The invention belongs to the technical field of microbial genetic engineering, and particularly relates to an acid-resistant lactobacillus plantarum engineering bacterium as well as a construction method and application thereof. The recombinant lactobacillus plantarum WCFS1-gadA-speA is obtained by constructing a recombinant expression vector pMG36e-gadA-speA and inducing the expression of two key genes, and the recombinant lactobacillus plantarum WCFS1-gadA-speA is obtained. The analysis of the fermentation capacity of the strain finds that compared with a single gene heterologous expression strain, the recombinant lactobacillus plantarum WCFS1-gadA-speA shows higher acid resistance and apple juice fermentation capacity. The recombinant lactobacillus plantarum WCFS1-gadA-speA with higher strong acid tolerance is obtained through a heterologous expression technology, a strain resource with better performance is provided for the food fermentation industry, and the quality and stability of a fermented product are improved.

The present invention discloses a recombinant saccharomyces cerevisiae for efficiently biosynthesizing inositol and its construction method and application, and belongs to the field of microbial genetic engineering. The recombinant saccharomyces cerevisiae of the present invention strengthens the expression of hexokinase gene hxk, inositol 3-phosphate synthase gene ino, inositol monophosphatase gene suhb, and realizes the rearrangement of glucose metabolism by knocking out the transcriptional repressor opi1 of inositol 3-phosphate synthase (Ino1) and glucose 6-phosphate dehydrogenase gene zwf1, and relieves feedback inhibition; In addition, the present invention further increases the yield of inositol by screening inositol external emission protein gene ompf, ompc and knocking out inositol internal transport protein gene itr1, itr2. The present invention realizes the efficient production of inositol (yield is as high as 195.7g / L), with good industrialization prospect.

The invention belongs to the field of microbial genetic engineering, and particularly relates to an efficient homologous recombination method suitable for bifidobacteria and application of the efficient homologous recombination method. Specifically, the invention discloses a method for constructing and screening the combination of the single-chain annealing protein and the single-chain binding protein for the first time. According to the method, a single-chain annealing protein and a single-chain binding protein are jointly constructed in an escherichia coli-bifidobacterium shuttle expression vector, and co-expression of the single-chain annealing protein and the single-chain binding protein in bifidobacterium longum is realized, so that the homologous recombination capability of the bifidobacterium longum is remarkably improved. According to the method disclosed by the invention, the homologous recombination efficiency of genome editing of the bifidobacterium longum is greatly improved, the absolute genome editing efficiency of the bifidobacterium longum is improved to 0.00009792% from the original level which is almost undetectable, and the research and application progress of genome engineering of the strain is remarkably promoted, so that the method has a good practical application value.

This invention belongs to the field of microbial genetic engineering technology, specifically relating to an α-2,6-sialic acid transferase mutant and its application. This invention utilizes a mutant derived from *Lithocarpus lucida* (…). Photobacterium Five amino acids in the amino acid sequence of α-2,6-sialyl transferase encoded by the gene of sp. JT-ISH-224 were mutated at specific sites to obtain α-2,6-sialyl transferase mutants M133L, E140D, D228H, N289D, and G322Q. Among them, the E140D and G322Q single-site mutants of α-2,6-sialyl transferase showed increases in 6'-SL yield of 87.80% and 78.01%, respectively, compared with wild-type α-2,6-sialyl transferase. The optimal recombinant microbial strain 6'-SL-7 (E140D / G322Q) obtained by combining the mutation sites achieved a yield of 1.471 g / L, effectively solving the bottleneck problem of low catalytic efficiency of sialyl transferase and providing a high-performance enzyme element for the efficient and low-cost biomanufacturing of 6'-SL.

This invention discloses a method for preparing self-healing concrete based on microbial gene programming, belonging to the field of building materials technology. The method includes: constructing a recombinant engineered strain incorporating a urease gene cluster and a carbonic anhydrase gene; preparing a spore suspension and protecting it with a three-layer microencapsulation system of sodium alginate / chitosan / gelatin; preparing nutrient-releasing microspheres encapsulating calcium lactate and urea in a polylactic acid-glycolic acid copolymer; and mixing cement, standard sand, water, fly ash, microencapsulated spore particles, nutrient-releasing microspheres, nano-silica, and modified bentonite to prepare self-healing concrete. This invention enhances the urease activity of the strain through genetic engineering, protects the spores using a three-layer microencapsulation technology, and combines a nutrient-releasing system and nano-synergists to achieve efficient self-repair of concrete cracks.

The present invention provides a microorganism fluorescence detection system and detection method, which relates to the field of microorganism gene detection technology. The detection system includes a body, a detection hole, a drive plate, a mounting plate, a holding bottle, a catheter and an automatic dripping mechanism. The detection hole is arranged on one side of the body, the drive plate is installed inside the detection hole, the mounting plate is installed on the body, and a plurality of holding bottles are installed on one side of the mounting plate, and one end of the catheter is connected to the holding bottle. The present invention can automatically control the liquid transfer gun to drip the liquid to be detected into multiple detection cavities of the detection dish. The automatic dripping process makes the experimental process more coherent and systematic, reduces the operation steps and time waste, improves the detection efficiency, reduces the error caused by human operation, and improves the reliability of the experimental results; and during the dripping process, the conveyor belt can seal and shield the detection cavity, thereby avoiding the detection liquid from sputtering during the dripping process, thereby facilitating the influence of sample loss and avoiding experimental environmental pollution.

This invention discloses a high-yield lactyl-3-4 ...5-4-4-4-4-4- N This invention relates to plasmid-free genetically engineered strains of tetrasaccharides and their applications, belonging to the field of microbial genetic engineering. The invention screened strains with higher activity. β -1,3-galactosyltransferase gene PmgalT Computer-aided enzyme function modification was used to achieve a shake-flask yield of 10.05 g / L for the constructed mutant. Subsequently, genome integration was performed... breast Multi-copy integration PmgalT * V27I / V39I / Y147F ‑galE‑galU A plasmid-free strain was constructed, enabling it to grow in shake flasks with lactoyl- N The yield of tetrasaccharides reached 14.4 g / L. Using glucose as the sole carbon source, the highest yield of 68.19 g / L was achieved at 64.5 h; using glycerol as the sole carbon source, the highest yield of 62.88 g / L was achieved at 60.5 h, providing a basis for its industrial production.

The invention relates to the field of microbial genetic engineering and synthetic biology, and particularly discloses a yersinia pseudotuberculosis temperature-sensitive promoter and application thereof. The screened and identified temperature-sensitive promoter element fills the blank of efficient constitutive promoter resources of yersinia pseudotuberculosis. The promoters can be used as universal gene expression regulation modules to be applied to metabolic engineering and synthetic biology research of yersinia pseudotuberculosis and other prokaryotic hosts, and are beneficial to high-efficiency expression of target protein or biosynthesis of high-value products. According to the method, the expression of the target gene can be regulated and controlled by changing the culture temperature without adding chemical inducers, so that the operation is convenient, the cost is saved, and the method has outstanding advantages in large-scale fermentation production.

The invention provides application of protein in construction of engineering bacteria for producing 6 '-sialic acid lactose, and relates to the technical field of microbial genetic engineering. According to the invention, new alpha2, 6-sialyltransferase with the potential of producing 6 '-sialyllactose is obtained through screening, and genes for coding precursor CMP-Neu5Ac synthetic pathway enzyme and alpha2, 6-sialyltransferase are introduced into escherichia coli to construct a new biosynthesis pathway of 6'-sialyllactose so as to construct engineering bacteria for efficiently producing 6 '-sialyllactose. Besides, a precursor CMP-Neu5Ac synthetic pathway enzyme and alpha2, 6-sialyltransferase are integrated and expressed in the engineering bacteria, so that the instability problem caused by plasmids is avoided, meanwhile, the dependence on antibiotics in the production process is reduced, and the environmental risk in the production process is reduced.

The invention discloses a knockout fragment of a biofilm dispersion regulating gene spo0J and its application, and belongs to the technical field of microbial genetic engineering. The knockout fragment of the biofilm dispersion regulating gene spo0J of the invention has a nucleotide sequence as shown in SEQ ID NO.3. The target fragment of the knockout spo0J gene, i.e., the knockout fragment of the biofilm dispersion regulating gene spo0J, is obtained by overlapping PCR; the fragment is transformed into a wild-type FZB42 by a chemical transformation method to obtain a spo0J knockout strain △spo0J. Compared with the wild-type FZB42, the △spo0J strain has a significantly improved level of indoleacetic acid (IAA) secretion; the relative expression levels of tryptophan synthesis genes trpB, trpC and indoleacetic acid (IAA) synthesis gene ysnE are significantly increased.

The present invention discloses a knockout system and application of a biofilm dispersion regulating gene spoIIIJ, and belongs to the technical field of microbial genetic engineering. The present invention obtains a biofilm regulating gene spoIIIJ by reverse PCR identification, and its nucleotide sequence is shown in SEQ ID NO.1; obtains a target fragment of the knockout spoIIIJ gene by overlapping PCR, that is, a knockout fragment of the biofilm dispersion regulating gene spoIIIJ; and transforms the fragment into a wild-type FZB42 by a chemical transformation method to obtain a spoIIIJ knockout strain △spoIIIJ. Compared with the wild-type FZB42, the △spoIIIJ strain has a significantly improved level of indoleacetic acid (IAA) secretion; the relative expression levels of tryptophan synthesis genes trpB, trpC and indoleacetic acid (IAA) synthesis gene ysnE are significantly increased.

The application belongs to the technical field of kidney disease differential diagnosis, and particularly relates to a microbial gene marker for differentiating diabetic nephropathy and membranous nephropathy and application thereof. The application provides sufficient evidence for the differentiation and treatment of clinical diagnosis of diabetic nephropathy and pathological diagnosis of membranous nephropathy based on four microbial markers of Blautia, Akkermansia, Sphingomonas and Granulicatella.

The invention discloses a method for increasing the yield of lactoyl-N-neotetraose based on genome integration and transporter screening, and belongs to the field of microbial genetic engineering. According to the invention, a lactoyl-N-trisaccharide II synthesis module and a lactoyl-N-neotetraose synthesis module are rapidly integrated to different sites of an escherichia coli genome in a multi-copy manner by virtue of a CRISPR (clustered regularly interspaced short palindromic repeats)-associated transsposases gene editing technology, so that the use of plasmids is reduced, and the metabolic burden of cells is reduced. And in a 3L bioreactor, the yield of the lactyl-N-neotetraose reaches 30.56 g / L. Through over-expression of the transporter, the transporter which is most obvious in improving the yield of the lactoyl-N-neotetraose is screened to be YbSET, the yield of the lactoyl-N-neotetraose reaches 54.12 g / L, and excellent industrial application prospects are shown.

The invention belongs to the technical field of biological medicines, and particularly relates to an oral microbial gene marker for noninvasive diagnosis of esophageal cancer patients and application of the oral microbial gene marker. The invention provides an oral microbial gene marker for distinguishing esophageal cancer patients from healthy people, which consists of six microbial genes as shown in SEQ ID NO: 1-6, and the microbial genes are enriched in the oral cavity of a human body. The microbial gene distinguishing model disclosed by the invention has good distinguishing ability in esophageal cancer patients and healthy control people, and the feasibility, applicability and universality of the microbial gene distinguishing model in the esophageal cancer patients are proved.