254 results about "Corynebacterium glutamicum" patented technology

The invention discloses a spermidine synthetase mutant and application of the spermidine synthetase mutant in synthesis of spermidine, and belongs to the technical field of genetic engineering and metabolic engineering. According to the method, the spermidine synthetase is mutated firstly, the efficiency of the spermidine synthetase is improved, and on the basis, the dissolved oxygen and fed-batch concentration are optimally controlled through fermentation, so that the yield of spermidine is improved. The corynebacterium glutamicum provided by the invention is an engineering bacterium with plasmids and without defects, and after the corynebacterium glutamicum is subjected to fed-batch fermentation in a 5L fermentation tank for 68 hours, the yield of spermidine reaches 25.1 g / L, and the sugar-acid conversion rate is 17.9%.

The present application relates to the technical field of biocatalysis and biosynthesis, and specifically discloses a natural blue pigment and a biosynthesis method therefor. The present application uses metabolically engineered bacteria to express indigoidine synthetase and phosphopantetheinyl transferase, to catalyze glutamine and N-acetylglutamine to biosynthesize a natural blue pigment N-acetylindigoidine, and deduces its molecular structure by means of mass spectrometry, nuclear magnetic resonance and other methods. The present application achieves catalytic synthesis of N-acetylindigoidine from glutamine and N-acetylglutamine in Escherichia coli, Corynebacterium glutamicum, Saccharomyces cerevisiae and Streptomyces. Compared with indigoidine, N-acetylindigoidine has a maximum absorption wavelength of 584 nm, possesses better color brightness, and has more stable coloration that is not easily faded, with a very broad application range and industrial production application prospects.

The present application relates to the technical field of biocatalysis and biosynthesis, and specifically discloses a blue pigment and a biosynthesis method thereof. In the present application, an indigoidine synthetase and a 4′-phosphopantetheinyl transferase are expressed by a metabolically engineered strain to catalyze the biosynthesis of the blue pigment N-acetyl-indigoidine from glutamine and N-acetylglutamine, and a molecular structure of the blue pigment is inferred by mass spectrometry, nuclear magnetic resonance spectroscopy, etc. The present application achieves the catalytic synthesis of N-acetyl-indigoidine from glutamine and N-acetylglutamine in Escherichia coli (E. coli), Corynebacterium glutamicum (C. glutamicum), Saccharomyces cerevisiae (S. cerevisiae), and Streptomyces. Compared with indigoidine, N-acetyl-indigoidine has a maximum absorption wavelength of 584 nm, and a stable color having high brightness that is not easy to fade. Thus, the blue pigment shows an extensive application range and a promising industrial production prospect.

The invention belongs to the technical field of synthetic biology, and particularly relates to recombinant escherichia coli for efficiently producing O-acetyl-L-homoserine as well as a construction method and application of the recombinant escherichia coli. According to the invention, CRISPR-Cas9 gene editing is combined with metabolic modification, so as to precisely integrate OAH biosynthesis key nodes: introducing and overexpressing a mutant metX gene at a genome pseudogene site to enhance the expression of homoserine acetyltransferase, knocking out byproducts such as poxB and mgsA to synthesize genes, and carrying out high-yield synthesis of OAH. Corynebacterium glutamicum thrA (anti-feedback inhibition), aspB and bacillus subtilis acsA-acuA genes (construction of an acetic acid switch) are introduced, gene expression of gltA, tpiA and the like is optimized to increase supply of precursors such as acetyl CoA and the like, and finally a stable strain without plasmids, antibiotics and inducers is obtained. The production cost is effectively reduced, and the method has the advantage of high genetic stability and shows a good application prospect in OAH industrial production.

The invention discloses a production strain for fermenting L-histidine and application of the production strain, and belongs to the technical field of biological engineering. According to the invention, escherichia coli E.coli CICC 10243 is taken as an original strain, histidine synthesis operon genes hisDCB-cg2302-cg2301-HA-impA-FI, hisN and hisEG from corynebacterium glutamicum are integrated on a genome of the escherichia coli E.coli CICC 10243, a pyruvate dehydrogenase AceE mutant coding gene is introduced, and an escherichia coli transcription factor SlyA coding gene slyA is inactivated. The novel strain E.coli His08 for fermentation production of L-histidine is prepared and fermented in a 5L fermentation tank for 60 h, the histidine yield reaches 58.39 g / L, the sugar-acid conversion rate is 0.15 g / g, and the strain E.coli His08 has important application value.

The invention provides a beta-alanine producing strain as well as a construction method and application thereof. According to the bacterial strain, an acetaldehyde dehydrogenase gene adhE, a lactic dehydrogenase gene ldhA, an acetokinase gene ackA, a pyruvate dehydrogenase gene poxB, a DNA binding transcription inhibition factor coding gene lacI, an aspartate kinase thrA, a pantothenic acid synthase gene panC, an alanine synthesis transaminase gene cycA and an alanine synthesis transaminase gene yfbQ are knocked out from an E.coli W3110 genome, and the bacterial strain is obtained. A phosphoenolpyruvate carboxylase gene ppc, a pyridine nucleotide transhydrogenase coding gene pntAB, an aspartic acid transaminase gene aspC, an aspartic acid decarboxylase gene panD derived from pseudomonas aeruginosa and a beta-alanine transporter gene NCgl0580 derived from corynebacterium glutamicum are integrated at the same time, and the bacillus subtilis is used for producing beta-alanine and has the advantage of high fermentation yield.

The invention provides an L-arginine production strain and a construction method and application thereof, corynebacterium glutamicum AJC is modified by a metabolic engineering method to obtain the L-arginine production strain, the synthesis path of arginine is optimized, and a carbon source efficiently flows to arginine by means of promoter replacement, multi-copy and the like; by introducing an exogenous gene pyrABE949 *, the supply of a precursor substance carbamyl phosphate is enhanced; by introducing an exogenous gene pntAB, a new direction is provided for supply of coenzyme NADPH, and a large amount of reducing power is provided for synthesis of arginine; an exogenous gene fxpk is introduced, an NOG system is constructed, an acetyl coenzyme A pool is enriched, a large number of acetyl coenzyme A precursors are provided, and the engineering bacterium has the characteristics of high yield and stability and has good industrial application value in the aspect of fermentation production of L-arginine.

A C. glutamicum strain has enhanced productivity of L-lysine, and a method produces L-lysine using the C. glutamicum strain. The C. glutamicum strain expresses NAD(P)(+) transhydrogenase subunit alpha PntA and NAD(P)(+) transhydrogenase subunit beta PntB combined with increased activity of the myo-inositol transporter IolT. A method produces L-lysine in a fermentative process employing such C. glutamicum strain.

The embodiment of the invention discloses a recombinant III-type humanized collagen as well as a preparation method and application thereof. The amino acid sequence of the recombinant III-type humanized collagen is shown as SEQ ID NO.1, SEQ ID NO.2 or SEQ ID NO.3. The recombinant III-type humanized collagen has the advantages that the recombinant III-type humanized collagen can be used for preparing the recombinant III-type humanized collagen; the recombinant III-type humanized collagen provided by the invention has the activity of promoting cell adhesion, the amino acid sequence of the recombinant III-type humanized collagen is selected from the amino acid sequence of natural collagen, and the recombinant III-type humanized collagen does not generate immune response when being applied to a human body. The characteristic that corynebacterium glutamicum does not secrete endotoxin is utilized, the preparation method is simple, the product can meet the requirements of food, cosmetics, medical instruments and drugs without complex separation, and reference is provided for low-cost industrial production.

Provided are a theanine-producing strain and use thereof in tea fermentation production. A corynebacterium glutamicum is proposed, which includes an alanine decarboxylase CsAlaDC mutant. The theanine-producing strain is obtained by taking the corynebacterium glutamicum as a starting strain, knocking out in sequence an α-ketoglutarate dehydrogenase E1 subunit gene odhA, a glutamate external transporter gene Ncg11221 and a lactate dehydrogenase gene ldh; and / or expressesing a citrate synthase gene gltA, a pyruvate kinase gene pyk and a glutamate dehydrogenase gene gdh; and / or overexpressing an alanine dehydrogenase alaA and integrating a γ-glutamine synthetase GMAS into a cg1960 pseudogene locus of the corynebacterium glutamicum.

The application discloses an Escherichia coli probiotic with high beta-alanine yield and application thereof, and the recombinant genetically engineered bacterium takes a wild-type Escherichia coli probiotic as a starting strain and is subjected to one or more gene edits, including Trc promoter strengthening of a panD gene, an aspC gene, an sdhC gene and a ppc gene, overexpression of a panD gene from Bacillus subtilis, an aspB gene from Corynebacterium glutamicum, an aspA gene and a pyc gene from Corynebacterium glutamicum on a plasmid pTrc99a, and knockout of a cycA gene. 600 Compared with an existing reported Escherichia coli probiotic beta-alanine production strain, the OD of the recombinant genetically engineered bacterium is 4.4 times that of the existing reported strain, and the beta-alanine yield is 6 times that of the existing reported strain, so that the recombinant genetically engineered bacterium has great industrial application potential.

The invention discloses a corynebacterium glutamicum engineering strain produced by L-glutamine as well as a construction method and application of the corynebacterium glutamicum engineering strain. The corynebacterium glutamicum engineering strain is prepared by integrating a glutamine synthase encoding gene glnA which is sourced from saccharomyces cerevisiae and has a tyrosine residue mutation at the 99th site on the surface and an sRNA-MicC-EcHfq regulation module of a targeted adenylyltransferase gene glnE into a vector plasmid; the method comprises the following steps: constructing a recombinant plasmid for removing adenosine acylation of glutamine synthase; and transferring the recombinant plasmid into corynebacterium glutamicum of which the odhA gene expression is controlled by a growth stage specific promoter Pcg2705, so as to obtain a corynebacterium glutamicum engineering strain for producing L-glutamine. According to the invention, through multi-gene synergistic modification and space-time metabolic flux optimization, the problem of'growth-synthesis' tradeoff in the prior art is solved, the fermentation efficiency is improved, the fermentation process is simple, and the application prospect is good.

The invention relates to the technical field of synthesis of natural dyes by a biological method, in particular to an antibiotic-demand-free ornamental blue high-yield strain as well as construction and application of the antibiotic-demand-free ornamental blue high-yield strain. Corynebacterium glutamicum is used as an initial strain, a tricarboxylic acid circulating gene is regulated and interfered, and the copy number of exogenous blue pigment synthetase BpsA in a genome is adjusted, so that the ornamental blue high-yield strain is obtained. The strain does not need to use antibiotics in the fermentation process, the yield is stable, the product production capacity equivalent to the level when antibiotics are used can still be achieved through fermentation under the antibiotic-free condition, and the growth performance of the strain is kept. The method for producing the ornamental blue by using the strain provided by the invention ensures that the ornamental blue product is natural and green, also improves the efficient production capacity of the strain, solves the contradiction between green environmental protection and stable high yield of the ornamental blue dye, is green and environment-friendly, and has a remarkable industrial application prospect.

The invention discloses a rigorous induction system for corynebacterium glutamicum and application of the rigorous induction system, and belongs to the fields of genetic engineering and microbiology. The system can be strictly regulated when an inducer is not added, and can express only after the inducer is added. More specifically, the invention relates to the reduction of leak expression of Cre recombinase by comprising a logic gate, repression regulation and RNA regulation elements, and the use thereof for mediating SCRaMbLE to produce large-scale genome random rearrangement in Corynebacterium glutamicum synthetic genomes. Particularly, the method is used for reducing the leakage expression level of Cre recombinase and keeping the moderate expression level after induction. More importantly, the rigorous induction system disclosed by the invention can be applied to genome rearrangement to obtain rearranged strains with highly diversified genotype sequences, so that the purpose of increasing the yield of target products or enhancing the stress resistance of the strains is achieved.

The invention relates to the field of synthetic biology, metabolic engineering and fermentation engineering, and discloses a genetically engineered bacterium and application thereof in production of alpha-ketoglutaric acid.The genetically engineered bacterium comprises the steps that the genetically engineered strain with escherichia coli BW25113 or corynebacterium glutamicum ATCC13032 as a host is constructed; overexpression type isocitrate dehydrogenase genes are integrated through chromosomes, alpha-ketoglutarate dehydrogenase complex genes are knocked out, NADPH invertase is overexpressed through plasmids, and exogenous AKG transporter protein is introduced, so that metabolic flux directional regulation and control and product efflux enhancement are realized; meanwhile, a staged fermentation process is adopted and comprises dynamic gradient adjustment of inorganic phosphorus concentration and two-stage pH control. According to the method, the AKG yield of 78.2 g / L can be obtained under the 5L tank scale, the sugar acid conversion rate reaches 0.68, and the method has the advantages of being high in yield, low in byproduct and controllable in cost. The obtained AKG is suitable for a plurality of fields such as medical intermediates, functional foods and cell culture additives, and has wide industrial application prospects.

The invention discloses a corynebacterium glutamicum whole genome random mutation tool and application thereof, and belongs to the technical field of genetic engineering and biological mutagenesis. DsDNA is unwound in the transcription process based on DNA helicase, DNA adenine deaminase and the DNA helicase are fused, and whole-genome random mutation of corynebacterium glutamicum is achieved. The whole genome random mutation method disclosed by the invention is applied to mutagenesis of an original strain of Corynebacterium glutamicum ATCC 13032, and a mutant strain of which the biomass is 1.95 times of that of WT (OD600 is 2.2) and 1.99 times of that of WT / pXMJ19 (OD600 is 2.16) when being cultured under the condition that the pH value is 5.5 is obtained. The corynebacterium glutamicum whole genome random mutation method disclosed by the invention has important guiding significance for screening other mutation strains with enhanced robustness.

The invention provides a strain for producing L-2-aminobutyric acid as well as a construction method and application of the strain. According to the strain, E.coli W3110 is taken as a chassis strain, tdh, rhtA and ilvE genes are knocked out, the transcriptional levels of ppc, aspC, thrAfbrBC, ilvAfbr, tdcB, yaS and pntAB genes are up-regulated, an ilvI gene is weakened, bcd and gdh genes derived from bacillus subtilis B.subtilis 168 and a cysK gene derived from Corynebacterium glutamicum K051 are introduced in a heterologous manner, and the L-2-aminobutyric acid producing strain is obtained through screening and screening. The strain does not contain plasmids, is stable in heredity, does not need induction, does not need to add resistant substances, efficiently synthesizes L-2-aminobutyric acid from the beginning by taking glucose as a substrate, and has good L-2-aminobutyric acid synthesis capability, stable performance and high acid production efficiency.

The invention discloses a screening and expression platform construction method for non-essential gene loci of corynebacterium glutamicum, and belongs to the technical field of biology. The center sites CGLRS02940, CGLRS03085, CGLRS08555, CGLRS09020 and CGLRS13560 of the corynebacterium glutamicum identified by the invention have the remarkable advantages that when gene knockout or knockin operation is carried out, the sites do not interfere with the normal physiological metabolism process of strains, and stable expression of exogenous genes can be guaranteed; meanwhile, the gene editing efficiency is relatively high, and an editing target with high universality and high reliability is provided for genetic engineering modification of corynebacterium glutamicum.

The present invention relates to a Corynebacterium glutamicum mutant strain having increased L-citrulline productivity, and a method of producing L-citrulline using the same. The Corynebacterium glutamicum mutant strain is capable of producing L-citrulline in high yield and high concentration while inhibiting the production of by-products, because the activity of the protein that is expressed by the NCgl2657 gene therein has been weakened or inactivated.

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 technical field of biochemical engineering, and provides a method for improving the high-temperature tolerance of corynebacterium glutamicum and the production capacity of L-tryptophan. The invention provides a novel high-temperature tolerance related gene target osrR of corynebacterium glutamicum, biomass accumulation of corynebacterium glutamicum under a high-temperature condition is remarkably improved by regulating and controlling the expression level of the novel high-temperature tolerance related gene target osrR, and the fermentation yield of L-tryptophan of recombinant corynebacterium glutamicum under the high-temperature condition is improved. Specifically, by reducing the expression level of the recombinant corynebacterium glutamicum, the biomass accumulation of the recombinant corynebacterium glutamicum under the condition of 40 DEG C and the yield of the L-tryptophan under the condition of 33.5 DEG C are remarkably improved, and support is provided for high-temperature fermentation production of the L-tryptophan. Wide application prospects are realized.

The present invention discloses a method for producing creatine by fermenting saccharide raw materials using microorganisms, and belongs to the field of genetic engineering technology. L-arginine-glycine amidinotransferase and guanidineacetic acid N-methyltransferase with high enzyme activity are overexpressed in Corynebacterium glutamicum var.CCTCC AB 2021051 using the pXMJ19 plasmid to construct a creatine synthesis pathway. The degradation of SAH is promoted by integrating the expression of the SAHase enzyme, thereby relieving the inhibitory effect of SAH on the key enzymes in creatine production and promoting the regeneration of SAH substrate SAM. The engineered bacteria are scaled up in a 5L tank, fermented for 60 hours, and the creatine yield reaches 14.2g / L. The production method of creatine by the present invention has the advantages of low production cost and high efficiency, which is conducive to scaled-up production.

The invention discloses a method for non-induction biosynthesis of low molecular weight gamma-polyglutamic acid, which specifically comprises the following steps: cloning a gamma-polyglutamic acid synthetase gene cluster pgsBCA of bacillus licheniformis to corynebacterium glutamicum F343 with high glutamic acid yield for exogenous expression; a gamma-polyglutamic acid degrading enzyme gene pgdS and pgsBCA from Schlaughua Zehnder bacillus are subjected to tandem expression, so that the biosynthesis of the low-molecular-weight gamma-polyglutamic acid (LMW-gamma-PGA) is realized. Furthermore, natural promoters are screened, constitutive promoters with different intensities are introduced, and the optimal promoter Pdep-A16 is used for replacing the original inducible promoter Ptac, so that synthesis of the LMW-gamma-PGA without an inducer is realized. Finally, continuous fed-batch fermentation is carried out in a 15-L fermentation tank, the yield of the LMW-gamma-PGA reaches 37.42 g / L, and the production intensity is 0.78 g / L / h. And the LMW-gamma-PGA production process without adding an inducer is successfully constructed.

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.

This invention relates to a pyridoxal phosphate-responsive biosensor and its application, belonging to the field of synthetic biology biosensor technology. This invention is the first to discover and identify the relationship between pyridoxal phosphate and Cgl1185-P. cgl2838 The transcriptional regulation and response relationship between these factors was investigated, and this relationship was used as the basis for constructing a biosensor responsive to pyridoxal. The transcription factor regulatory system Cgl1185-P derived from Corynebacterium glutamicum was used. cgl2838 A biosensor for the specific detection of pyridoxal phosphate was constructed by assembling a reporter gene as the main component onto a plasmid, enabling effective detection of pyridoxal phosphate concentrations in intracellular or culture environments. This biosensor possesses advantages such as clear structure, high sensitivity, and good host adaptability, and can be used for fermentation process monitoring, cofactor metabolism research, and screening of relevant strains, showing promising application prospects in the fields of cofactor concentration monitoring and metabolic regulation.

The application discloses a construction method and application of a genetically engineered corynebacterium glutamicum for synthesizing a bacteriochlorophyllide amino acid shinorine, and belongs to the field of genetic engineering and fermentation technology. The genetically engineered corynebacterium glutamicum is constructed by taking corynebacterium glutamicum ATCC 13032 as a host strain, knocking out a competitive pathway gene, integrating a key enzyme gene in multiple copies on a genome, knocking out an acid production pathway gene and a serine dehydrogenase gene, and expressing a gene cluster for synthesizing shinorine on a plasmid, so that the synthesis yield of shinorine in a microbial cell is improved. The strain can produce shinorine in a flask, and 1.12 g / L of shinorine can be accumulated in 72 hours. In fed-batch fermentation in a 50 L fermenter, the strain can be cultured to 8.6 g / L.

The invention relates to Corynebacterium glutamicum with high indigo yield as well as a construction method and application thereof, and belongs to the technical field of gene recombination fermentation. The recombinant corynebacterium glutamicum can be used for overexpression of the following genes: a D-glucose-6-phosphate dehydrogenase gene zwf, a II type deoxyarabinose type heptulose phosphate synthase gene aroG and a flavin-containing monooxygenase gene CcFMO. The recombinant corynebacterium glutamicum can weaken the following genes: a pyruvate kinase gene pyk and a tryptophan synthase gene trpB. The corynebacterium glutamicum is used for overexpressing a zwf gene, an aroG gene and a CcFMO gene, weakening a pyk gene and a trpB gene and enhancing the expression quantity of an indole synthetic pathway related enzyme gene, so that the yield of the corynebacterium glutamicum at a shake flask stage reaches 619mg / L in a fermentation production process of indigo under the condition of not additionally adding tryptophan. The invention provides the recombinant corynebacterium glutamicum with high indigo yield, which is low in production cost, good in stability and high in synthesis efficiency, and is suitable for practical popularization and application.

The invention discloses a method for fermenting vinasse feed through mixed bacteria, and belongs to the field of microbial fermentation. The invention firstly provides a microbial agent for increasing the content of crude protein in fermented vinasse. The microbial agent contains a bacillus subtilis liquid, a corynebacterium glutamicum liquid, a saccharomyces cerevisiae liquid, an enterococcus faecalis liquid and a bifidobacterium animalis liquid. The invention also provides an application of the microbial agent in preparation of a fermented vinasse feed. The contents of crude protein, crude fat and amino acid in the fermented vinasse feed are increased. Through mixed fermentation, the content of amino acid in the vinasse is increased, the content of lysine is increased by 108.33%, and the content of threonine is increased by 59.09%.

The invention discloses a genetically engineered bacterium for efficiently producing D-pantothenic acid as well as a construction method and application of the genetically engineered bacterium, the construction method comprises the following steps: (a) taking corynebacterium glutamicum DPA2 as a chassis bacterium, enhancing the expression of at least one pantothenic acid main pathway gene in a chassis bacterium genome, and introducing plasmid pEC-XK99E-panBCilvC to obtain an engineered strain for modifying the pantothenic acid main pathway gene; (b) knocking out at least one heteroacid pathway gene in the engineering strain obtained in the step (a), carrying out in-situ integration on a panBCC gene at an alaT gene locus, and introducing a plasmid pEC-XK99E-panBCilvC to obtain a heteroacid pathway modified engineering strain; and (c) knocking out the pyruvic acid shunt gene pyc in the heteroacid pathway modified strain obtained in the step (b), enhancing the expression of an odx gene, and introducing a plasmid pEC-XK99E-panBCilvC to obtain the engineering strain for regulating and controlling the pyruvic acid shunt. The genetically engineered bacterium has the beneficial effects that the genetically engineered bacterium for efficiently producing D-pantothenic acid is obtained, and the yield and the conversion rate are higher; the level of the modified strain in the aspect of fermentation production of D-pantothenic acid is obviously improved.