21 results about "Yeast gene" patented technology

The present invention belongs to the field of bioengineering, genetic engineering technology and oligosaccharide (rare sugar) synthesis technology, and specifically relates to a method for producing lactose-N-disaccharide using genetically modified recombinant yeast. The technical scheme of the present invention is: a recombinant yeast cell is introduced with (1) a heterologous lactose-N-disaccharide phosphorylase ( LNBP ) gene, and (2) fructose-1-phosphate phosphorylase ( YQAB ) gene, and further preferably, also includes (3) destruction or modification of a marker gene. The recombinant yeast cell has the ability to synthesize lactose-N-biose (Lacto-N-biose, LNB) from lactose, and the yield of LNB is greatly improved by a molecular switch.

The invention provides auxotroph hansenula polymorpha as well as a construction method and application thereof. The method comprises the following steps: constructing a CRISPR gene editing plasmid of single-plasmid double-point editing yeast, introducing the plasmid into a hansenula polymorpha strain, and constructing the auxotroph hansenula polymorpha by replacing an sgRNA sequence in the plasmid, so as to obtain the auxotroph hansenula polymorpha. The auxotroph hansenula polymorpha keeps a high growth rate of an original strain, genetically engineered bacteria capable of efficiently producing exogenous target proteins can be rapidly screened out through auxotroph screening, expression products can be selectively accumulated in cells and can also be selectively secreted into fermentation liquor, and the yield of the heterotroph hansenula polymorpha is improved. The expressed recombinant protein has high biological safety and can be applied to multiple fields of medicine, medical beauty, instruments, food, animal husbandry and the like.

A genetically modified yeast cell, wherein the yeast cell comprises a genetic modification comprising overexpression of yeast gene encoding porphobilinogen deaminase (HEM3), the HEM3 gene having at least 80% identity with SEQ ID No. 7. The genome of the modified yeast cell further comprises one or more genetic modifications in one or more genes selected from: genes coding for heme-dependent repressor of hypoxic genes (ROX1), genes coding for heme oxygenase (HMX1), genes coding for a receptor for vacuolar proteases (VPS10), and genes coding for vacuolar proteinase (PEP4), the one or more genetic modifications being such that expression of a polypeptide from such a gene is reduced or disrupted or the polypeptide expressed is non-functional.

The invention discloses a method for improving yeast CRISPR-Cas gene editing efficiency, and belongs to the technical field of gene engineering. Firstly, SPC plasmids are transferred into host bacteria, the SPC plasmids contain gRNA1 expression cassettes targeting target genes 1, 5'ends of the gRNA1 expression cassettes are connected with Checkpoint promoters, the gRNA1 expression cassettes serving as host cells are transferred into pCasT plasmids, the plasmids and gRNA2 targeting yeast genome target genes 2 achieve DNA double-strand breakage through Cas protein, the Checkpoint promoters of the SPC plasmids are activated, transcription of the gRNA1 is started, and gene editing of the target genes 1 is achieved. On the contrary, the Cas protein and the gRNA1 expression cassette do not realize gene editing on the target gene 1, the unedited strain dies by absorbing the externally added toxic substances, and the successfully edited strain survives due to the lack of the functions. According to the method, the gene editing efficiency of a single target site can be improved, the gene knockout, large-fragment DNA insertion and recombination efficiency can be improved, and the high-throughput genome editing efficiency is improved.

The present application relates to a recombinant engineering bacterium producing human albumin and its construction method and application, belonging to the field of genetic engineering technology. In the present application scheme, the human albumin gene sequence is codon-optimized to construct a highly efficient recombinant human albumin Pichia yeast strain, further overexpress PDI and SSB2, and the gene sequence of the overexpressed protein in the recombinant plasmid is codon-optimized, and after transformation into the recombinant human albumin Pichia yeast strain, a Pichia yeast genetic engineering strain with high expression of human albumin is successfully obtained, and the strain achieves efficient expression of human albumin through high-density fermentation, with a yield of up to 38.6 g / L, laying the foundation for the industrial application of recombinant human albumin, and having important industrial application prospects and economic value.

The invention relates to a yeast gene modified cell containing GDP-fucose synthase polypeptide, which can be applied to synthesis of human milk oligosaccharide, and belongs to the technical field of gene engineering. According to the technical scheme, the yeast gene modified cell contains a heterologous nucleic acid sequence of a GDP-fucose synthase active polypeptide. The synthetase active polypeptide is obtained by modifying at least one site of the 20th site to the 29th site, the 40th site to the 53th site, the 75th site to the 81st site, the 95th site, the 148th site to the 158th site, the 193th site, the 215th site, the 219th site to the 21st site, the 227th site to the 232 site, the 239th site to the 241st site, the 259th site to the 264th site, the 301st site to the 302 site, the 306th site and the 310th site to the 316th site of an amino acid sequence shown in SEQ ID NO: 1. Compared with an original strain, the yeast gene modified cell has higher activity of synthesizing fucosyllactose, and a safe and feasible technical scheme is provided for industrial production of fucosyllactose.

Lipid production control factors in yeasts are provided. A yeast 115694 gene, a yeast 4888 gene, and a yeast 45559 gene are provided as lipid production control factors.

The invention discloses kluyveromyces marxianus genetically engineered bacteria and application thereof, and belongs to the technical field of bioengineering. The invention discloses a kluyveromyces marxianus genetically engineered bacterium. The kluyveromyces marxianus genetically engineered bacterium is prepared by the following steps: firstly, screening to obtain a kluyveromyces marxianus strain with high ergosterol yield; on the basis, inducing and separating a MATa haploid strain; then, through the introduction of heterologous DHCR24 and DWF5 genes, the weakening of a competitive pathway gene ERG6, the improvement of the metabolic flux of a squalene pathway in a sterol synthesis process, the compartment engineering of a peroxisome region of a mevalonic acid pathway, the knockout of an hypoxia inhibition transcription factor ROX1, the improvement of cofactor supply, the optimization of a yeast lipid pathway and other metabolic engineering optimization reconstructions, the expression of the yeast lipid pathway is improved; and an engineering strain with high cholesterol yield is constructed and obtained. Through fed-batch fermentation, efficient and stable synthesis of cholesterol is achieved, the yield reaches 7.64 g / L, and a foundation is laid for biological manufacturing of cholesterol.

This invention discloses a fully humanized N-glycosylated yeast strain and its applications. The modification of the yeast strain in this invention includes the knockout of basal N-glycosylation, the construction of a humanized N-glycosylation pathway, the integration of genes related to heparin precursor synthesis, and the integration of genes for heparin-modifying enzymes. The N-glycosylated Pichia pastoris platform strain described in this invention represents the first time that full humanization has been achieved, with its N-glycosyl group including fucosylate. When expressing animal-derived heparin-modifying enzymes, the fully humanized Pichia pastoris platform strain of this invention exhibits increased enzyme activity, thereby increasing the sulfate groups in the heparin glycan chains, thus enhancing the biological activity of heparin.

The present invention provides a yeast genetic engineering bacterium for producing resveratrol and a construction method and application thereof. The engineering bacterium overexpresses resveratrol synthase on the chromosome of Saccharomyces cerevisiae BY4742. STS , p-Coumaryl-CoA ligase 4CL , phenylalanine / tyrosine ammonia lyase TAL , cinnamate-4-hydroxylase C4H , cytochrome P450 enzyme reductase CPR1 , DAHP synthase mutant ARO4 , chorismate mutase mutant ARO7 Chorismate synthase ARO2 shikimate kinase AroL , acetyl-CoA synthetase ACS and malonyl-CoA synthetase ACC1 , and knock out bypass genes CIT2, LPP1, DPP1, and PDC6 at the same time, to achieve biosynthesis of resveratrol using glucose as a substrate. The present invention utilizes phenylalanine / tyrosine ammonia lyase to react with phenylalanine and tyrosine as substrates to generate key precursors for resveratrol synthesis, strengthens the precursors required for resveratrol synthesis, and greatly increases the yield of the product resveratrol.

The invention relates to the technical field of gene engineering, in particular to a yeast gene engineering strain for producing parthenolide and a construction method of the yeast gene engineering strain. According to the present invention, it is determined that the improvement of the heme supply in the yeast gene engineering strain for producing parthenolide is beneficial to the improvement of the parthenolide yield, and it is proved that the integration site of the heme synthesis gene has effects on the parthenolide yield, such that the powerful evidence and the basis are provided for the subsequent continuous optimization and parthenolide yield increasing.

Lipid production control factors in yeasts are provided. A yeast 115694 gene, a yeast 4888 gene, and a yeast 45559 gene are provided as lipid production control factors.

This invention discloses a genetically engineered strain of *Kluyveromyces martensii* and its applications, belonging to the field of bioengineering technology. The invention discloses a genetically engineered *Kluyveromyces martensii* strain, firstly obtained through screening a strain that produces high levels of ergosterol; based on this, it induces and isolates... Eye A haploid strain was constructed and then metabolically optimized through the introduction of heterologous DHCR24 and DWF5 genes, weakening of the competitive pathway gene ERG6, increasing the metabolic flux of the post-squalene pathway during sterol synthesis, peroxisome compartmentalization engineering of the mevalonate pathway, knockout of the hypoxia-inhibiting transcription factor ROX1, improved cofactor supply, and optimization of the yeast lipid pathway, resulting in a high-cholesterol-producing engineered strain. Through fed-batch fermentation, efficient and stable cholesterol synthesis was achieved, with a yield of 7.64 g / L, laying the foundation for cholesterol biomanufacturing.

A yeast cell for the production of fatty acids and / or fatty acid-derived products is genetically modified for overexpression of one or more endogenous yeast genes selected from the group consisting of ADP-ribosylation factor-binding protein, phosphatidylinositol 4,5-bisphosphate 5-phosphatase INP54, NADP-dependent isocitrate dehydrogenase IDP3, mitochondrial phosphatidylglycerophosphatase GEP4, triglyceride lipase TGL1, long chain fatty acyl-CoA synthetase FAA3, lipid phosphate phosphatase LPP1, and variants of such endogenous yeast genes. The yeast cell produces higher amount of fatty acids and / or fatty acid-derived products compared to a yeast cell not overexpressing the one or more endogenous yeast genes.

The application discloses a recombinant Pichia pastoris genetic engineering bacterium with high yield of lactoferrin, and simultaneously satisfies the following three modifications: overexpression of a bovine lactoferrin expression gene, mutation of an alpha mating factor signal peptide at position E86 to F, and overexpression of a PDI molecular chaperone gene. The application also discloses a construction method of the recombinant Pichia pastoris genetic engineering bacterium, and application of the recombinant Pichia pastoris genetic engineering bacterium in production of bovine lactoferrin. The 5L fermenter test result of the bacterium strain shows that the yield of exogenous protein is significantly improved, and is increased from 10.04 mg / L in a flask fermentation to 95.78 mg / L, nearly 10 times. The application is based on the Pichia pastoris system and is not related to complex culture condition change, and is low in cost and easy to enlarge. The method is not only suitable for production of bovine lactoferrin, but also provides a new modification idea for high-efficiency expression of other heterologous proteins in the yeast system.

The invention discloses application of saccharomyces cerevisiae and metabolites thereof in preparation of anti-aging products. The saccharomyces cerevisiae is screened out from a saccharomyces cerevisiae single-gene mutant strain library, and the saccharomyces cerevisiae lacks one of genes with nucleotide sequences shown as SEQ ID NO.1-14. The invention further discloses a preparation method of the saccharomyces cerevisiae. The yeast gene engineering strain and the metabolite thereof have the functions of remarkably prolonging the life, improving the exercise ability and the intestinal integrity of an old person and delaying the aging of the body; in addition, standardized production of the strain can be realized.

The invention discloses pichia pastoris genetically engineered bacteria based on enzyme efficient surface display of adhesion protein, a construction method of the pichia pastoris genetically engineered bacteria and application of the pichia pastoris genetically engineered bacteria in immobilized continuous catalysis. An adhesion protein gene, a beta-galactosidase gene LacA and a cell wall anchoring protein gene Pir1p are heterologously expressed in starting strain pichia pastoris. According to the pichia pastoris genetically engineered bacterium, cell wall adhesion protein genes are integrated and expressed in a pichia pastoris GS115 genome, and the cell film-forming capacity of pichia pastoris is remarkably enhanced; and performing multi-copy integration on the beta-galactosidase surface display gene, and controlling the target protein beta-galactosidase to be displayed on the surface of the yeast by using a constitutive promoter. The pichia pastoris genetically engineered bacterium based on the high-efficiency surface display of the enzyme of the adhesion protein can be used for multi-batch continuous catalysis, the effect is better than that of the enzyme surface display genetically engineered bacterium which is not integrated with a biological membrane gene, the relative conversion rate is still kept at 78% after six batches, and the pichia pastoris genetically engineered bacterium has a good industrial application prospect in immobilized continuous catalysis.

The present invention discloses a Cas9 mutant for improving gene editing efficiency and its application, relating to the field of gene editing technology. The amino acid sequence of the Cas9 mutant is shown in SEQ ID NO.17. This method can be used not only for gene editing in Saccharomyces cerevisiae, but also for other microorganisms that use this gene editing principle. The present invention can not only be used to study the function and metabolic mechanism of yeast genes, but also the mutant strains obtained do not contain any residual foreign genes and can be safely used in industrial production.

The present invention relates to a yeast gene ScFIT3 for regulating and controlling cadmium resistance of an organism and an application thereof. The yeast gene ScFIT3 for regulating and controlling the cadmium resistance of the organism has a nucleotide sequence as shown in SEQ ID NO.1. The present invention discloses, for the first time, the application of the yeast gene ScFIT3 in enhancing cadmium resistance of the organism. The yeast gene ScFIT3 is linked to an expression vector and transformed into a budding yeast, thereby completing construction of a cadmium-resistant yeast. The growth of the budding yeast transformed with the yeast gene ScFIT3 is significantly superior to that of a wild-type budding yeast, indicating that the budding yeast transformed with the yeast gene ScFIT3 has good resistance to cadmium stress, significantly enhances a capability of resisting heavy metal cadmium, and reduces accumulation and absorption of cadmium.

The present invention relates to a yeast gene ScFIT3 for regulating the cadmium resistance of organisms and its application. The nucleotide sequence of the yeast gene ScFIT3 for regulating the cadmium resistance of organisms is shown in SEQ ID NO.1. The present invention discloses for the first time the application of the yeast gene ScFIT3 in enhancing the cadmium resistance of organisms. The yeast gene ScFIT3 is connected to an expression vector and transformed into budding yeast to complete the construction of cadmium-resistant yeast. The growth of the budding yeast transformed with the yeast gene ScFIT3 is significantly better than that of the wild-type budding yeast, indicating that the budding yeast transformed with the yeast gene ScFIT3 has excellent resistance to cadmium stress, significantly enhanced resistance to heavy metal cadmium, and reduced cadmium accumulation and cadmium absorption.

The invention discloses a yeast genetically engineered bacterium for enhancing secretory expression of heterologous protein and application of the yeast genetically engineered bacterium. According to the invention, yeast is used as an original strain, the yeast genetically engineered bacterium is constructed by introducing genes, and the introduced genes comprise any one of the following groups: (1) grpE gene; (2) a groES gene; (3) a groEL gene; (4) a PDI1 gene; (5) an ERO1 gene; (6) a dnaJ gene; (7) a tig gene; and (8) a dnaK gene. The chaperonin widely used in escherichia coli is applied to pichia pastoris for the first time, so that the secretory expression efficiency of heterologous protein is remarkably improved. According to the invention, glycosyl transferase EUGT11m which is difficult to express in pichia pastoris is selected as a model protein, and experiments find that after the molecular chaperones are introduced into the pichia pastoris, the EUGT11m not only realizes effective secretory expression, but also confirms the functionality of the EUGT11m through enzyme activity detection.