20 results about "Fed-batch culture" 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 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 application discloses a kind of high-yield O-succinyl-L-homoserine recombinant escherichia coli and its construction method and application, belong to genetic engineering and fermentation engineering technical field.The recombinant escherichia coli is expressed by expressing anti-feedback inhibition homoserine O-succinyltransferase metA fbr , aspartate ammonia-lyase aspA, aspartate transaminase aspC, 2 ketoglutarate decarboxylase sucA, dihydrothioctic acid succinyltransferase sucB and efflux protein yjeH, knock out metJ, improve intracellular NADPH availability, to enhance OSH synthesis pathway metabolic flow, precursor succinyl coenzyme A supply and product efflux.Using 5 L bioreactor for fed-batch fermentation, after fermentation 60 h, OSH production is as high as 137.4 g / L, and sugar acid conversion rate reaches 50.5%.The application provides engineering strain and technical realization path for the efficient fermentation of OSH.

This invention discloses a high-yielding erythritol-producing *Yarrowia lipophila* strain (… Yarrowia lipolytica This invention relates to the field of biotechnology, specifically to *Yarrowia lipophila* CA20 as the starting strain, and its applications. A000039 A gene knockout method was used to construct a high-yield erythritol-producing *Yarrowia lipolytica* strain. The construction method provided by this invention is simple and of significant value in simplifying the construction process of high-yield strains. Furthermore, under the fermentation medium and conditions provided by this invention, the *Yarrowia lipolytica* strain producing high-yield erythritol was fed-batch fermented with glucose as a substrate, achieving a yield of 280.50 g / L and a yield of 0.71 g / g, demonstrating good economic benefits.

The invention provides a construction method and application of recombinant yarrowia lipolytica with high yield of salidroside, and belongs to the field of bioengineering. The recombinant yarrowia lipolytica with high yield of salidroside is named as a yarrowia lipolytica XJ-SAL strain, and after the recombinant yarrowia lipolytica XJ-SAL strain is subjected to fed-batch fermentation, the salidroside can be synthesized at a relatively high level. The construction method of the recombinant yarrowia lipolytica is easy to operate, and efficient synthesis of salidroside can be achieved.

The invention provides a construction method and application of recombinant yarrowia lipolytica with high hydroxytyrosol yield, and belongs to the field of bioengineering. The recombinant Yarrowia lipolytica capable of producing the hydroxytyrosol at the high yield is named as a Yarrowia lipolytica XJ-HT strain, and after the recombinant Yarrowia lipolytica XJ-HT strain is subjected to fed-batch fermentation, the hydroxytyrosol can be synthesized at a relatively high level. The construction method of the recombinant yarrowia lipolytica is easy to operate, and efficient synthesis of hydroxytyrosol can be achieved.

The application discloses a construction method and application of Corynebacterium glutamicum for synthesizing shinorine, and belongs to the field of genetic engineering and fermentation technology. The Corynebacterium glutamicum ATCC 13032 is used as a starting strain, a competitive pathway gene is knocked out, an optimal exogenous enzyme is screened, and a key enzyme gene cluster is integrated in multiple copies on a genome, so that the synthesis yield of shinorine in the microbial cell is improved, and the strain can accumulate 0.63 g / L of shinorine in 72 hours in a flask. In fed-batch fermentation in a 50 L fermenter, the final yield of shinorine can reach 4.3 g / L.

The invention discloses an engineering strain for increasing supply of cofactors and application of the engineering strain in improving production of epsilon-polylysine, and belongs to the technical field of microbial genetic engineering. The method comprises the following steps: by taking streptomyces albus for producing epsilon-polylysine as an original strain, simultaneously connecting a nicotinic acid ribose phosphate transferase gene pncB, a phosphoenolpyruvate carboxylase gene pck and a pyruvate dehydrogenase gene pdh in the strain with a pIB139 vector by utilizing a homologous recombination method to obtain a recombinant plasmid vector pIB139-pncB-pck-pdh, and introducing the recombinant plasmid vector into a competent cell to obtain the recombinant plasmid vector pIB139-pncB-pck-pdh. The engineering bacterium BP3 is obtained. The shake flask epsilon-PL yield of the engineering bacterium BP3 reaches 1.61 g / L and is improved by 30.89% compared with that of an original strain; the yield of fed batch fermentation reaches 37.21 g / L, the yield reaches 4.85 g / (L.d), and the maximum thallus concentration DCW reaches 43.47 g / L.

Improved production of recombinant proteins in E. coli, reliant on tightly controlled autoinduction, triggered by phosphate depletion in stationary phase. The process also provides an optimized autoinduction media, enabling routine batch production at various culture volumes where cells densities routinely reach ˜5-7 g cell dry weight per liter and offer protein titers above 2 g / L. The methodology has been validated with a set of diverse heterologous proteins and is of general use for the facile optimization of routine protein expression from high throughput screens to fed-batch fermentation.

The invention belongs to the technical field of epsilon-polylysine production, and particularly relates to a screening culture method and application of tolerant streptomyces hygroscopicus. The screening culture method of the tolerant streptomyces hygroscopicus comprises the following steps: subculturing streptomyces hygroscopicus engineering bacteria in a culture medium taking straw cellulose hydrolysate as a carbon source for at least 30 generations to obtain primary streptomyces hygroscopicus. The primary streptomyces hygroscopicus is subcultured for 10-25 generations in a fermentation culture medium with the epsilon-polylysine concentration being 0.5 g / L-5g / L and straw cellulose hydrolysate as a carbon source, and the tolerant streptomyces hygroscopicus is obtained. Wherein the tolerance means that epsilon-polylysine is produced while an inhibitor is resisted. The straw cellulose hydrolysate contains an inhibitor. According to the screening culture method of the tolerant streptomyces hygroscopicus, disclosed by the invention, the yield of epsilon-polylysine in virus-free cellulose hydrolysate can be up to 1.35 g / L at most, and the yield of epsilon-polylysine can be up to 30.57 g / L at most under the condition of fed-batch fermentation in a 5L fermentation tank.

The invention belongs to the technical field of biological medicines, provides an efficient screening method for a high-yield carotenoid rhodosporidium toruloides space mutant strain, and belongs to the technical field of biological medicines, aiming at the problems that existing carotenoid high-yield strain screening depends on color observation, the positive mutation rate is low, the screening workload is large and the like. According to the core scheme, a YPD solid basic culture medium and a compound growth inhibitor (containing D-glucose, D-xylose, vanillin and other components) are mixed according to a certain volume ratio to prepare a screening culture medium, and the space mutation rhodosporidium toruloides mutant strain for high-yield carotenoid is efficiently screened out. Furthermore, the screened high-yield carotenoid rhodosporidium torulosporidium space mutant strain is subjected to fed-batch fermentation to produce carotenoid, the yield can reach 48.7 mg / L, and the method has a good application prospect.

Disclosed herein are methods, cells, and compositions for the production of bacterial cell extracts. The bacterial cell extracts are used in cell-free protein synthesis of recombinant proteins and large quantities of bacterial cell extracts are typically needed. The present disclosure relates to the production of high cell density fermentations that provide large quantities of bacteria for the manufacturing of bacterial cell extracts. The present disclosure relates to high density fed batch fermentation for the production of bacterial cell extract. Also provided herein are cell-free protein synthesis systems and methods that use the bacterial cell extracts.

The invention provides recombinant yarrowia lipolytica with high grease yield and a construction method of the recombinant yarrowia lipolytica, and belongs to the field of bioengineering. The Yarrowia lipolytica strain po1g is used as an original strain to integrally express the diacylglycerol acyltransferase from the cocoa tree; an acetyl-coenzyme A carboxylase of yarrowia lipolytica; a human source stearoyl coenzyme A deunsaturated enzyme; a yarrowia lipolytica AMP deaminase; the aldehyde dehydrogenase of the escherichia coli obtains a recombinant strain for improving the yield of the grease of the yarrowia lipolytica. Preferably, the screened recombinant yarrowia lipolytica strain with high grease yield is named as the yarrowia lipolytica strain GK03, 131 g / L of fatty acid can be produced after fed-batch fermentation at a 5L tank level, and efficient synthesis of the fatty acid in the yarrowia lipolytica is realized. The method provided by the invention can be used for constructing the high-oil-yield yeast, and has wide application prospects in the fields of renewable energy sources, chemical industry, medicine and health care and the like.

The application discloses a kind of production phycobilisome amino acid Porphyra-334 genetically engineered glutamic acid coryneform bacteria and application thereof, belong to microbial genetic engineering and fermentation technical field.The application uses glutamic acid coryneform bacteria ATCC 13032 as host strain, by knocking out competitive pathway gene, in genome multiple copy source from Nostoc linckia NIES-25 for synthesizing Porphyra-334 four key synthesis pathway enzyme genes, knocking out acid-producing pathway gene and threonine dehydrase gene in strain to realize the improvement of synthesis Porphyra-334 yield.Finally, genetically engineered strain produces Porphyra-334 in shake flask, and can accumulate 1.23 g / L in 72 h.In 50L fermenter, fed-batch fermentation is expanded in culture, and finally can reach 7.08 g / L.

The invention provides a genetically engineered bacterium for synthesizing vanillyl alcohol by taking glucose as a substrate and application of the genetically engineered bacterium, the genetically engineered bacterium comprises a protocatechuic acid synthesis module and a vanillyl alcohol synthesis module, the protocatechuic acid synthesis module expresses a 3-dehydroshikimic acid dehydratase gene, and the vanillyl alcohol synthesis module expresses a 3-dehydroshikimic acid dehydratase gene. The vanillyl alcohol synthesis module is used for expressing a carboxylic acid reductase gene, a phosphopantetheinyl transferase gene and a 3-O-methyltransferase gene. According to the genetically engineered bacterium constructed by the invention, glucose is used as a substrate to synthesize vanillyl alcohol, so that the synthesis step of vanillyl alcohol is shortened, no additional substrate is needed, the yield of vanillyl alcohol obtained by shake flask fermentation can reach 1145.94 mg / L, and the yield of vanillyl alcohol obtained by fed-batch fermentation can reach 1007.07 mg / L.

The invention discloses a cytochrome oxidase P450 BM3 mutant and application thereof, and belongs to the technical field of bioengineering. According to the invention, the cytochrome oxidase P450 BM3 derived from bacillus megatherium is subjected to site-directed mutagenesis to obtain the cytochrome oxidase P450 BM3 mutant with obviously improved catalytic activity and selectivity, and when the 2-phenethyl alcohol derivative is prepared, the P450 BM3 mutant can obviously improve the yield of the 2-phenethyl alcohol derivative; furthermore, cells for efficiently synthesizing the 2-phenethyl alcohol derivative from the beginning by taking glucose as a substrate are constructed, the high yield of the 2-phenethyl alcohol derivative is realized, the highest yield of the (R)-phenethyl alcohol derivative is 3.54 g / L, and the final yield is increased to 12.4 g / L through fed-batch fermentation of a 5L fermentation tank, which is the highest yield of the (R)-phenethyl alcohol produced by known microorganisms at present; and the yield of the salidroside reaches 546.20 mg / L.

The application discloses a mutagenic high-yield acid sophorolipid Torulopsis globosa and a fermentation culture method. Starmerella bombicola The mutagenic high-yield acid sophorolipid Torulopsis globosa (Torulopsis globosa) strain G72 is obtained by using Torulopsis globosa ATCC 22214 as a starting strain, compound mutagenesis of UV and EMS and grading screening under high-sugar conditions, and is preserved in the Guangdong Microbial Digital Museum Center with a preservation number of GDMCC 67888. The application further provides a segmented fed-batch fermentation process for synergistically controlling high-sugar and low-oil of the mutant strain G72, through segmentally regulating a sugar-oil ratio, pH and dissolved oxygen, the end-point concentration of sophorolipid can be stabilized at about 347.94 g / L, the acid sophorolipid concentration is 267.25 g / L, the proportion is more than 80%, the fermentation period is shortened, and the high selectivity, high space-time yield and low-cost production of the acid sophorolipid are realized.

The invention relates to a method for reinforcing crosslinking of an escherichia coli outer membrane and peptidoglycan to promote biosynthesis of beta-caryophyllene, in the biosynthesis process of terpenoid substances, the terpenoid compounds damage the integrity of cell membranes mainly by destroying the stability of the outer membrane, and in order to overcome the challenge, the biosynthesis of the beta-caryophyllene is promoted. A novel'cell membrane damage defense model 'is generated at the right time. By enhancing crosslinking between an outer membrane and peptidoglycan, the stability of the membrane is maintained under the stress of terpenoids, so that the metabolic activity of cells is remarkably improved. Through strategies of optimizing metabolic pathways, enhancing cell viability, refining fermentation process regulation and control and the like, the yield of the beta-caryophyllene reaches 15.6 g / L after 49 hours of fed-batch fermentation, and the highest record of producing the beta-caryophyllene by escherichia coli within the shortest fermentation period is created. The research provides a new method for effectively synthesizing other terpenoids to solve the challenge of cell membrane damage.

This invention relates to the field of biotechnology, and more particularly to recombinant strains producing nicotinamide mononucleotide (NMN) and their applications. This invention enhances NMN accumulation through metabolic flux redirection and gene knockout, successfully establishing an NMN synthetic pathway in *E. coli*. Furthermore, by optimizing the NAM-NMN transport system and the PRPP synthesis pathway, the NMN yield is effectively increased to 7.0 g / L. This invention also optimizes the NMN production process, achieving a maximum NMN yield of 28.5 g / L in a fed-batch fermentation system, a significant increase compared to traditional batch fermentation.

The invention discloses a construction method and application of a saccharomyces cerevisiae recombinant strain, a key gene of a carnitine shuttle transport pathway is screened out by adopting the carnitine shuttle transport pathway and regulating and controlling the carnitine shuttle transport pathway, and the yield of hypotanshinone diene is further improved; a high-yield tanshinone diene mutant M30 is screened out by integrating key genes in a synthetic route of the tanshinone diene in multiple copies; through 5L fed batch fermentation culture, the yield of the hypotanshinone diene is 3.7 g / L, which is reported so far, the highest yield of the hypotanshinone diene synthesized from the beginning of the saccharomyces cerevisiae, a biosynthesis pathway of the hypotanshinone diene is successfully constructed in the saccharomyces cerevisiae, the synthesis efficiency of the hypotanshinone diene in the saccharomyces cerevisiae is remarkably improved, and the yield of the hypotanshinone diene in the saccharomyces cerevisiae is greatly improved. And the method has important significance for realizing the production of other high-added-value abietane type diterpenoid compounds.