431 results about "Metabolic engineering" patented technology

The invention discloses construction and application of an L-malic acid producing issatchenkia orientalis strain, and belongs to the technical field of microorganisms and the technical field of microbial fermentation and metabolic engineering. Through a metabolic engineering means, a cytoplasm metabolic pathway of an issatchenkia orientalis strain is transformed, and a pyruvate decarboxylase coding gene, glycerol dehydrogenase and lactic dehydrogenase are knocked out; a pyruvate carboxylase gene derived from aspergillus flavus, a malate dehydrogenase gene derived from corynebacterium glutamicum and phosphoenolpyruvate carboxylase derived from escherichia coli are screened and integrated, so that a cytoplasm rTCA cycle synthesis pathway is enhanced; inserting a dicarboxylic acid external transporter gene from schizosaccharomyces cerevisiae at a neutral site GD21, and knocking out endogenous dicarboxylic acid internal transporter; the L-malic acid yield of the modified yeast strain in the fermentation process is remarkably increased and reaches 140.16 g / L. The engineering strain provided by the invention has good industrial application potential in efficient L-malic acid production.

The invention relates to a construction method of escherichia coli engineering bacteria with high yield of N-acetylglucosamine. According to the method, the metabolic network of escherichia coli is directionally modified through metabolic engineering, the problems of'product re-decomposition ', by-product accumulation, poor genetic stability and the like of natural strains are solved, and efficient industrial production of GlcNAc is realized.

The invention discloses a rhodosporidium toruloides strain with high homologous recombination efficiency as well as a construction method and application thereof, and belongs to the field of oil-producing yeast engineering bacteria. According to the rhodosporidium toruloides strain, the rhodosporidium toruloides strain NP11-SaCas9-1 is used as an original strain, and an RAD52 gene is over-expressed. The modified yeast engineering bacterium provided by the invention can realize high homologous recombination through CRISPR system mediation, the homologous recombination efficiency reaches 90-100%, a multi-gene editing platform can be constructed on the basis, and the modified yeast engineering bacterium can be applied to the fields of synthetic biology, metabolic engineering, genetic engineering and the like, so that the modified yeast engineering bacterium is simpler, quicker, more convenient and more accurate in the genetic modification process. And the homologous recombination activity of the rhodosporidium toruloides can be improved by expressing the homologous recombination related protein Rad52, and the genetic engineering strategy can be used for modifying other unconventional yeasts.

The invention discloses a combined module for improving the yield of actinomycetes polyketone natural products and construction and application of recombinant bacteria containing the combined module, and belongs to the technical field of genetic engineering. In order to improve the yield of natural products such as polyketone, non-ribosome peptide and the like, the invention provides a combined module capable of improving the yield of actinomycetes natural products by combining and transforming a synthesis pathway of CoASH and a post-modification pathway of PPTase through a metabolic engineering strategy, and the module consists of CoASH cofactor synthesis module elements SCoaAR106A and SCoaD and a PPTase post-modification module element HPC3. By constructing a recombinant vector and recombinant bacteria containing the combined module, the yield of natural products such as actinomycetes polyketone, polyether and non-ribosomal peptide can be effectively improved by over-expression of the combined module.

The invention relates to an L-valine production strain as well as a construction method and application thereof, the strain is obtained by transformation through a metabolic synthesis path of L-valine, and the construction method comprises the following steps: by taking Cornebacterium glutamicum ATCC 13032 (C.glutamicum) as an original strain, carrying out editing transformation such as gene knockout, gene overexpression, gene heterologous expression, gene double-copy expression and the like on a genome of the original strain by utilizing a metabolic engineering means. The L-valine production strain disclosed by the invention can be used for synthesizing L-valine from the beginning by taking glucose as a carbon source, has the advantages of high production rate, short fermentation period, high strain stability and low cost, and lays a foundation for realizing large-scale production of L-valine. According to the fermentation tank fermentation method, the L-valine is produced in a mechanical stirring type fermentation tank by using the L-valine production strain Va1-09, 86.4 g / L of L-valine is produced by fermenting for 36 hours, and the method has a good industrial prospect.

The invention provides a genetically engineered bacterium for producing O-succinyl-L-homoserine as well as a construction method and application of the genetically engineered bacterium. In a chassis bacterium genome, the expression of a 2-ketoglutaric acid decarboxylase encoding gene sucA is enhanced, and the expression of a succinyl-coenzyme A synthetase encoding gene sucD is weakened, so that the supply of succinyl-coenzyme A is increased; the method comprises the following steps: increasing the NADPH (Nicotinamide Adenine Dinucleotide Phosphate) reducing capacity and ATP (Adenosine Triphosphate) energy supply of a chassis bacterium, increasing DNA (Deoxyribose Nucleic Acid) in combination with a transcription dual regulatory factor ompR to improve the stress resistance of escherichia coli under high osmotic pressure, and introducing an overexpression plasmid containing a homoserine transsuccinylase coding gene metA to construct the genetically engineered bacterium for producing O-succinyl-L-homoserine. The engineering strain obtained through a systematic metabolic engineering modification strategy can realize effective accumulation of OSH, the shake flask yield of OSH reaches 19.8 g / L, the fed-batch fermentation yield of a 5L fermentation tank reaches 110.5 g / L, the sugar-acid conversion rate reaches 52.6%, and a foundation is laid for subsequent construction of high-yield OSH engineering bacteria.

The present disclosure relates to the seminal discovery of a generation and use of genetically engineered Vibrio sp. Provided is the use of the genetically engineered bacteria for the construction, maintenance, manipulation, and / or propagation of DNA constructs; protein expression; protein secretion; vectors and other metabolic tools; metabolic engineering; expression of cellular extracts for cell-free biology; shuttle vectors; cloning vectors; and for synthetic biology applications. The disclosure also relates to the use of the replication machinery of Vibrio sp. as a cloning or expression vector for replication of recombinant DNA constructs. The disclosure also relates to methods of use of the above.

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 invention relates to the technical field of synthetic biology, metabolic engineering and cell culture. The present invention provides a cell for the production of a negatively charged, preferably sialylated, oligosaccharide wherein the cell is genetically engineered to have or express, preferably overexpress, a hydrolytic UDP-N-acetyl-D-glucosamine-2-epimerase. The invention further provides the use of said cells in culture or incubation. Also described are methods of producing negatively charged, preferably sialylated oligosaccharides using the cells, and purification of the negatively charged, preferably sialylated oligosaccharides.

The invention provides construction and application of a hyaluronic acid recombinant bacterium, and belongs to the fields of biochemical engineering, synthetic biology, genetic engineering and metabolic engineering, the recombinant corynebacterium glutamicum contains a hyaluronic acid synthase gene, a UDP-glucose dehydrogenase gene and a glutamide-6-phosphate fructose transaminase gene; the hyaluronic acid synthase gene, the UDP-glucose dehydrogenase gene and the glutamine-6-phosphate fructose transaminase gene are respectively connected with a sequence RBS1, a sequence RBS2 and a sequence RBS3; the recombinant bacterium provided by the invention also contains a dCas9 protein gene dCas9 and any one of genes sgRNA1, sgRNA2, sgRNA3, sgRNA4, sgRNA5 and sgRNA6, and the recombinant bacterium provided by the invention is high in hyaluronic acid yield and low in production cost.

The embodiment of the invention provides a recombinant pichia pastoris strain as well as a construction method and application thereof. The construction method comprises the following steps: by taking a pichia pastoris strain as a host strain, constructing a dual-synthesis path of phenylalanine ammonialyase and tyrosine ammonialyase in the host strain to obtain a first engineering strain; removing chorismic acid feedback inhibition of the first engineering strain based on metabolic engineering to obtain a second engineering strain; optimizing an aromatic amino acid synthesis route of the second engineering strain to obtain a third engineering strain; knocking out a branch metabolic pathway of the third engineering strain to obtain a fourth engineering strain; increasing precursor supply of the fourth engineering strain to obtain a recombinant pichia pastoris strain; according to the construction method provided by the embodiment of the invention, the synthesis path of p-coumaric acid can be systematically optimized, the recombinant pichia pastoris strain for efficiently producing p-coumaric acid by taking methanol as a carbon source is constructed, and compared with a host strain, the recombinant pichia pastoris strain has the advantage that the yield of p-coumaric acid is remarkably increased.

The invention relates to a method for producing glycosylglycerol. The method comprises the following steps: S1, culturing trichocephalus microcoleus to a logarithmic phase; and S2, adding glycerol and glucose into the culture environment of the coleus microcoleus to induce the coleus microcoleus to accumulate the glycosylglycerol. According to the invention, by optimizing the culture of the coleus microcoleus BL0902, the production of the glycosylglycerol by using the strain is realized, and on the basis, the strain is subjected to metabolic engineering transformation, so that the yield of the glycosylglycerol is greatly improved by the obtained engineering strain.

The invention discloses a yarrowia lipolytica engineering strain with high yield of resveratrol as well as construction and application of the yarrowia lipolytica engineering strain, and belongs to the technical field of synthetic biology and metabolic engineering. The invention provides a yarrowia lipolytica engineering strain. A 4-coumaric acid-CoA ligase gene, a multi-copy resveratrol synthase gene, a transketolase 1 gene, a fusion gene of a histidine phosphate aminotransferase gene and a tyrosine ammonia lyase gene, a glycolytic pathway hexokinase gene, a 6-phosphofructokinase-1 gene and a phosphoglycerate kinase gene are integrated, so that the recombinant protein is obtained. According to the present invention, by using glucose as the substrate, the high yield of resveratrol can be achieved by using glucose as the substrate, the resveratrol yield during shake-flask culture fermentation can achieve 3.157 g / L, the resveratrol yield can be increased to 30.7 g / L during 5L-scale fed-batch fermentation, and the resveratrol yield can be increased to 30.7 g / L when the resveratrol yield is increased to 30.7 g / L when the resveratrol yield is increased to 30.7 g / L when the resveratrol yield is increased to 30.7 g / L; the maximum value is reported by the yarrowia lipolytica system at present.

The invention belongs to the technical field of metabolic engineering and biological manufacturing, and particularly relates to an engineering bacterium for producing gallic acid (GA) and application of the engineering bacterium in production of gallic acid. According to the method, key metabolic nodes in the gallic acid production process are systematically regulated, for example, the hydroxylase activity is improved, the pyruvic acid flow direction is regulated, the aromatic amino acid precursor supply is improved, and the dynamic balance between the engineering bacterium growth and the gallic acid product synthesis is realized, so that the gallic acid yield and the carbon yield are remarkably improved. Experiments show that the gallic acid yield of the engineering bacteria prepared by the invention reaches 12.5 g / L in a shake flask experiment, and reaches 109.8 g / L in a 3 L fermentation tank; and the carbon yield is increased to 0.64 g / g glucose, which is the highest level reported in the current literature.

The invention relates to recombinant escherichia coli for de novo synthesis of piceatannol as well as a construction method and application thereof, and belongs to the field of synthetic biology and metabolic engineering. According to the invention, an L-tyrosine production strain is used as an original strain, five key genes required for synthesizing piceatannol are combined and connected to a plasmid vector, and a recombinant plasmid is introduced into the original strain, so that an engineering strain for heterologous synthesis of piceatannol is successfully constructed. According to the invention, the yield of piceatannol is further obviously improved through copy number optimization.

Escherichia coli is modified by means of metabolic engineering, a genetically engineered bacterium for high yield of O-succinyl-L-homoserine is obtained, and a construction method of the genetically engineered bacterium comprises the following steps: weakening a synthetic route of an organic acid metabolic byproduct of a chassis strain E.coli W3110 [delta] metI [delta] metJ [delta] thrB [delta] metB; wherein the synthetic route of the organic acid metabolic byproducts is selected from at least one of a formic acid synthetic route, a lactic acid synthetic route and an ethanol synthetic route. The constructed genetically engineered bacterium is applied to microbial fermentation production of the O-succinyl-L-homoserine, effective accumulation of the O-succinyl-L-homoserine is achieved, the sugar acid conversion rate reaches 46.8%, and a foundation is laid for subsequent construction of high-yield O-succinyl-L-homoserine engineering bacteria.

The invention discloses recombinant escherichia coli capable of increasing the yield of fucosyllactose and a construction method and application of a fucosyltransferase mutant of the recombinant escherichia coli, and belongs to the field of synthetic biology and metabolic engineering. According to the invention, recombinant escherichia coli B20 and B21 are constructed based on key enzyme engineering transformation. Under the shake flask condition, the 3-FL production capacity of the recombinant strain is obviously improved to 6.20 g / L and 6.34 g / L from the initial 3.45 g / L, and the amplification reaches 79.7% and 83.8% respectively. The yields of 3-FL of the strain B20 and the strain B21 respectively reach 53.88 g / L and 54.64 g / L under the culture condition of a 3L fermentation tank. The recombinant escherichia coli for expressing the 2 '-FL is constructed, and the yield of the 2'-FL reaches 64.86 g / L in a 3L fermentation tank. The invention provides a new technical scheme for efficient microbial synthesis of the 3-FL.

This disclosure is in the technical field of synthetic biology and metabolic engineering. More particularly, this disclosure is in the technical field of cultivation or fermentation of metabolically engineered cells. This disclosure describes a cell metabolically engineered for production of a mixture of at least three different oligosaccharides. Furthermore, this disclosure provides a method for the production of a mixture of at least three different oligosaccharides by a cell as well as the purification of at least one of the oligosaccharides from the cultivation.

The application provides a recombinant Pichia pastoris strain and a construction method and application thereof. The construction method comprises the following steps: taking a Pichia pastoris strain as a host strain, constructing a double synthesis pathway of phenylalanine ammonia lyase and tyrosine ammonia lyase in the host strain to obtain a first engineering strain; relieving the branch acid feedback inhibition of the first engineering strain based on metabolic engineering to obtain a second engineering strain; optimizing the aromatic amino acid synthesis pathway of the second engineering strain to obtain a third engineering strain; knocking out the branch metabolic pathway of the third engineering strain to obtain a fourth engineering strain; increasing the precursor supply of the fourth engineering strain to obtain the recombinant Pichia pastoris strain. The construction method can systematically optimize the synthesis path of p-coumaric acid, and the recombinant Pichia pastoris strain for efficiently producing p-coumaric acid with methanol as a carbon source is constructed. Compared with the host strain, the yield of p-coumaric acid of the recombinant Pichia pastoris strain is significantly improved.

The invention provides a strain for producing L-isoleucine as well as a construction method and application thereof. The strain is obtained by modifying a chassis strain escherichia coli XX12 by utilizing a metabolic engineering modification method, ldhA, adhE and pflB genes are deleted, the transcriptional levels of aspC, pykF, pntAB, ppK, ppc, ilvAfbr, ilvIHfbr and ygaZH genes are up-regulated, the transcriptional level of a leuA gene is down-regulated, a bcd gene derived from B.subtilis 168 and ppnK and cysK genes derived from Cornebacterium glutamicum ATCC 13032 are heterologously expressed, and the strain has the advantages that the strain can be used for producing L-isoleucine; the method has the advantages of no need of adding resistant substances, good L-isoleucine synthesis capability, short fermentation period by using glucose as a carbon source, and good economic benefit and industrial application value.

The invention discloses application of a gene GlZn2Cys661 in regulation and control of synthesis of ganoderic acid. The nucleotide sequence of the gene GlZn2Cys661 is as shown in SEQ ID NO. 1. The gene expression is regulated and controlled by constructing an overexpression or silent vector, so that the synthesis of the ganoderic acid is promoted or inhibited, and an effective method is provided for regulating and controlling the yield of the ganoderic acid. The invention relates to the technical field of bioengineering, and discloses an application of a gene GlZn2Cys661 in regulation and control of ganoderic acid synthesis. The invention provides a key gene target and a core engineering strain for efficiently producing ganoderic acid through a metabolic engineering means, and has important application value.

The invention belongs to the technical field of biology, and particularly discloses a high-density fermentation method and application of recombinant spider silk protein, and the high-density fermentation method comprises the following steps: firstly, transforming an expression vector carrying spider silk protein genes into modified pichia pastoris cells to obtain a strain; the activated strain is subjected to expanding culture, high-density fermentation culture and purification, and recombinant spider silk protein is obtained; wherein the transformation means of the pichia pastoris cell is to integrate Vitreoscilla hemoglobin Vhb gene into the pichia pastoris cell by utilizing a gene recombination technology, and the expression of the Vitreoscilla hemoglobin Vhb gene is improved by carrying out metabolic engineering transformation on the pichia pastoris cell for expression and optimizing the regulation and control of staged fermentation parameters, so that the expression efficiency of the Vitreoscilla hemoglobin Vhb gene is improved. The problems that in existing recombinant protein fermentation, oxygen dependence is high, product degradation is serious, purity is low, and large-scale stability is poor are solved, the yield and purity of recombinant spider silk protein are guaranteed, and large-scale production of the recombinant spider silk protein is facilitated.

The invention relates to the field of microbial metabolism engineering, and discloses a method for increasing the yield of 2 '-fucosyllactose (2'-FL) through the synergistic effect of various alpha-1, 2-fucosyltransferases. According to the method, two or more alpha-1, 2-fucosyltransferases with different sources are co-expressed in microbial cells, and the complementary enzymatic characteristics of the alpha-1, 2-fucosyltransferases are utilized, so that the synthesis efficiency of the 2 '-FL is remarkably improved, and the yield of the 2'-FL is effectively improved. The microbial cell is escherichia coli C43 (DE3) (E. coli C43 lacZ waaF with lacZ and waaF genes knocked out, the serial number is SL), a pETDuet-1 plasmid is constructed, and genes manB, manC, gmd, wcaG and zwf are overexpressed on a pETDuet-1 vector, so that the microbial cell can be used for preparing the microbial cell. In a 5L fermentation tank, after the final strain SL07 is used for final fermentation for 68 hours, the yield of 2 '-FL reaches 148g / L, and the production intensity reaches 2.18 g / L / h. The embodiment shows that the engineering strain for co-expressing the alpha-1, 2-fucosyltransferase from different sources can obviously improve the yield of the 2 '-FL.

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.

The invention relates to a strong promoter P7 suitable for streptomyces and application thereof, and relates to the field of genetic engineering and microbial metabolism engineering. The nucleotide sequence of the strong promoter P7 is as shown in SEQ ID No.1, and the strong promoter comprises a plasmid vector of the strong promoter; a host cell comprising the plasmid vector; the invention also discloses application of the strong promoter, the plasmid vector and the host cell in starting expression of a target gene. Compared with the prior art, the characterization of the strong promoter P7 provides an effective tool element for streptomyces strong promoter engineering and high-efficiency gene expression, and has important significance on streptomyces silent gene characterization, high-efficiency gene expression, metabolite synthesis, metabolic pathway reconstruction and the like. The strong promoter can be applied to common streptomyces type strains, and has important significance on high yield of important proteins including enzymes and important metabolites from actinomycetes.

The invention provides an oxazomycin high-yield engineering strain as well as a construction method and application thereof. The method for constructing the oxazomycin high-yield engineering strain comprises the following steps: driving high expression of a gene cluster by using a strong promoter in streptomyces longsheng, enhancing transcription of a key resistance gene module (ozmS-oxaA), and optimizing ribosome engineering of strain chassis cells and a culture medium. According to the invention, the rational synthetic biology and metabolic engineering technology is combined with the irrational ribosome engineering technology to gradually increase the yield of the streptomyces longsheng oxazomycin, and the established efficient biosynthesis system lays a foundation for further development of the oxazomycin.

The invention discloses pichia kudriavzevii for high yield of L-malic acid as well as a construction method and application thereof, and belongs to the technical field of biology, the pichia kudriavzevii is named as YM-023 and classified as Pichia kudriavzevii, the preservation number is CGMCC No. 35487, the preservation time is July 31, 2025, and the preservation number is CGMCC No. 35487. The preservation unit is China General Microbiological Culture Collection Center (CGMCC). The method comprises the following steps: starting from an acid-resistant yeast Kudriavzevii pichia pastoris strain YL-000 (the preservation number is CGMCC No.35680), carrying out metabolic engineering transformation to obtain a transformed strain, and ensuring that the transformed strain can realize efficient production of L-malic acid in a fermentation mode of adding a small amount of calcium carbonate neutralizer.

The invention belongs to the field of metabolic engineering, and discloses a genetically engineered bacterium for producing myo-inositol as well as a construction method and application of the genetically engineered bacterium. The genetically engineered bacterium takes escherichia coli as an original strain, and is obtained by performing the following gene editing on a genome of the escherichia coli: non-expressed lactose operon repressor protein, glucose phosphate dehydrogenase, acetokinase, glucose phosphate isomerase and glycerol repressor protein; and overexpressing inositol-1-phosphate synthase, inositol monophosphate, glucokinase, glucose permease and glycerol kinase. Compared with the prior art, the genetically engineered bacterium disclosed by the invention has remarkable advantages in the aspects of key enzyme expression rate, metabolism specificity, fermentation period and the like, and an efficient, stable, economical and feasible solution is provided for industrial production of myo-inositol.

The invention provides recombinant escherichia coli, a construction method of the recombinant escherichia coli and application of the recombinant escherichia coli in production of indoleacetic acid, and belongs to the technical field of synthetic biology and microbial metabolism engineering. The recombinant Escherichia coli is obtained by taking Escherichia coli as a chassis cell, introducing an IAA pathway into the chassis cell and knocking out a tnaA gene and an mtr gene; the recombinant escherichia coli can be fermented by taking glucose or cellulose as a substrate to generate IAA; the yield of the synthesized IAA is improved by further optimizing the fermentation time, the fermentation temperature and the composition of a buffer solution, and the synthesized IAA can effectively promote plant growth and has good practicability.

The invention belongs to the technical field of molecular biology, and particularly relates to a DNA binding site of a fungus transcription factor AniJ and application. According to the invention, an echinocandin B gene cluster regulatory factor AniJ is taken as a research object, a ptH-SpRY-ACBE base editor and an sgRNA library covering an aniA promoter are utilized, 5-FOA screening is combined, and a binding motif RNGCTGAS of the AniJ in a core promoter region is excavated. The construction of an engineering strain containing a modified promoter (increasing the number of motifs) proves that the motifs can significantly enhance the expression of aniA and the yield of echinocandin B. And a novel molecular tool is provided for metabolic engineering of natural products of fungi.