11 results about "Protease Gene" patented technology

The invention provides bacillus licheniformis for high-yield protein glutaminase as well as a construction method and application of the bacillus licheniformis, a target strain is obtained by knocking out an alkaline protease gene aprE and extracellular protease genes bpr, epr, vpr or wpr and heterologously expressing the protein glutaminase gene, an EPE strain is optimal in expression, PG enzyme activity reaches 0.358 U / mL and is improved by 3.55 times compared with that of a wild type, and the bacillus licheniformis has the advantages that the bacillus licheniformis is high in yield and high in protein glutaminase yield. In addition, the growth condition of each mutant strain is not obviously abnormal, and the growth speed is obviously increased. It is fully proved that the expression level of the heterologous protein PG can be remarkably improved by targeted knockout of the extracellular protease gene of the bacillus licheniformis, and a theoretical basis is laid for construction of an efficient protein expression host. The invention not only provides a new scheme for high-efficiency expression of PG, but also provides important reference for expression optimization of heterologous proteins of other gram-positive bacteria, and shows a wide industrial application prospect.

The application discloses a pichia pastoris bioengineering bacterium for relieving degradation of recombinant collagen, a preparation method and application thereof. Through bioinformatics analysis, gene knockout and function verification screening, three protease genes capable of relieving degradation of collagen in a pichia pastoris host are obtained, namely, chr1-4_0611, chr1-4_0362 and chr2-2_0380. Knocking out the protease chr1-4_0611 can make the total protein yield accumulation increase to 3.5 times of that of a control bacterium, and the proportion of a target band is greater than 50%, which can provide good theoretical and technical guidance for effective expression of other recombinant proteins in the pichia pastoris host.

The application belongs to the technical field of enzyme engineering, and particularly relates to a basic protease mutant and application thereof. The application provides a basic protease mutant, which is based on 2709 basic protease gene derived from bacillus licheniformis, and is obtained by random mutation through error-prone PCR and then screening. The basic protease obtained by the application can resist high-salt environment, can maintain good activity and enzymolysis efficiency in the high-salt environment, and after being treated in 20% (w / v) NaCl high-salt environment for 3 hours, the protease activity retention rate is increased by 25-50% compared with the original protease, the enzymolysis capacity in the 20% (w / v) NaCl high-salt environment is nearly 2-3 times that of the original protease, and the application can meet the application requirements in high-salt scenes such as food fermentation.

The invention discloses a preparation method of a protease overexpression yeast strain, the protease overexpression yeast strain and application of the protease overexpression yeast strain, and relates to the technical field of molecular biology.According to the technical scheme, a first expression plasmid of protease A is provided; carrying out one-step assembly on the first expression plasmid, a second expression plasmid containing the TPI1 gene and a third basic vector, and screening and verifying to obtain a third expression plasmid; the third expression plasmid is transferred into a yeast strain, a protease overexpression yeast strain is obtained through screening and verification, the yeast strain is a TPI1 gene knockout strain, a TPI1 gene transcription unit and a protease A gene transcription unit are connected in series and introduced into the yeast strain with the TPI1 gene knockout in a plasmid form, and overexpression of protease is achieved. The yeast genome does not need to be directly edited. The method not only simplifies the construction process, but also possibly improves the genetic stability of the engineering strain, and reduces the potential risk caused by genome integration.

This invention relates to the field of biotechnology, and discloses an engineered *Yarrowia lipolytica* strain that produces p-coumaric acid using glucose as a substrate, its construction method, and its applications. Construction method: A tyrosine ammonia-lyase gene is integrated using a CRISPR / Cas9 localization and integration method. TAL In *Yarrowia lipophila* strains, the DAHP synthase gene was enhanced. ARO4 , DHS1 and AROG Overexpression of tyrosine synthase TYR Genes and histidine phosphotransferases HIS5 The protease gene was further integrated to synthesize exogenous phenylalanine deamination and hydroxylation pathway genes for p-coumaric acid, including phenylalanine ammonia-lyase gene, cinnamate hydroxylase gene, and P450 reductase gene. The p-coumaric acid produced by the engineered *Yarrowia lipolytica* strain of this invention can reach a maximum yield of 1.7 g / L in shake flasks, and a yield of 30 g / L in a 5 L fed-batch fermentation tank, demonstrating significant industrial application value.

The invention belongs to the technical field of plant molecular biology, and discloses a soybean disease resistance-related protease GmRD21A gene, a coding protein thereof, and an application of the gene in plant disease resistance. The nucleotide sequence of the protease GmRD21A gene is as shown in SEQ ID NO. 1, and the amino acid sequence of the polypeptide coded by the protease GmRD21A gene is as shown in SEQ ID NO. 2. Protein spectrum analysis of phytophthora sojae pectate lyase PsPL1 proves that the protease GmRD21A degrades pectate lyase secreted by phytophthora and can resist attack of pathogenic bacteria pectate lyase on plant cell walls, and meanwhile, overexpression of the GmRD21A gene in soybeans can remarkably improve the resistance of plants to phytophthora. The invention can be applied to crop breeding anti-epidemic disease improvement, and is hopeful to improve the disease resistance of plants to epidemic diseases, thereby achieving the purposes of increasing yield and reducing pesticide consumption.

The invention relates to application of a protease gene in improving the yield of bacillus amyloliquefaciens iturin A and a construction method of bacillus amyloliquefaciens for high-yield iturin A, and belongs to the technical field of genetic engineering and microorganisms. Experiments prove that compared with wild bacillus amyloliquefaciens, the yield of the iturin A of the bacillus amyloliquefaciens of which the nucleotide sequence is deleted, such as a protease gene shown in SEQ ID NO.8, is remarkably increased by about 76%. Furthermore, on the basis that a protease gene with a nucleotide sequence as shown in SEQ ID NO.8 is deleted, the expression of a gene ndk related to ATP synthesis is enhanced, and it is found that compared with wild bacillus amyloliquefaciens, the yield of the iturin A in the constructed strain is remarkably increased, and the yield of the iturin A is remarkably increased. And compared with bacillus amyloliquefaciens which only lacks a nucleotide sequence, such as a protease gene as shown in SEQ ID NO.8, the bacillus amyloliquefaciens is also remarkably improved.

The present application belongs to the field of biomedical technology, and particularly relates to a method for screening of quorum sensing system inhibitors of Pseudomonas aeruginosa and simultaneously determining the target of the inhibitors. The method comprises the following steps: constructing a QS inhibitor screening system, and the QS inhibitor screening system takes protease as a screening marker; constructing a mutant with a deleted protease gene, which provides QS signal molecules in the process of co-culture; and screening a compound with inhibitory effect on the QS system and determining the target of the compound by means of co-culture or exogenous addition of QS signal molecules. The present application establishes a simple, fast, effective and high-throughput QSI screening method, and the target of the compound can be determined simultaneously.

The application belongs to the technical field of biology, and discloses application of mf1-01441 gene, protease and recombinant strain; specifically, application of mf1-01441 gene in preparation of protease and application of mf1-01441 gene in preparation of recombinant strain capable of producing protease, wherein the nucleotide sequence of the mf1-01441 gene is shown as SEQ ID NO. 1; the application successfully screens a novel protease mf1-01441 gene by constructing a Fosmid library of intestinal tract microorganisms of gibel carp; the library not only breaks through the limitation of traditional culture method, captures genetic information of a large number of uncultured microorganisms in the intestinal tract of gibel carp, lays a foundation for exploring microbial resources in the unique habitat, but also provides high-quality materials for screening of functional genes such as protease, and is expected to provide a new idea for improving feed utilization rate of fish and reducing breeding cost, and promote sustainable development of green breeding.

The present application belongs to the technical field of genetic engineering and enzyme engineering, and particularly relates to a low-temperature neutral protease from marine microorganism colwellia psychrerythraea, a preparation method thereof and application of the low-temperature neutral protease in hydrolysis of proteins. In the present application, a protease gene from colwellia psychrerythraea is connected with an expression vector pPIC9K to obtain a recombinant plasmid, then the recombinant plasmid is electrotransformed into Pichia pastoris cells, and a target gene is expressed by using methanol induction, and a centrifugal product is obtained after removing supernatant, thereby obtaining a recombinant low-temperature neutral protease. The recombinant low-temperature neutral protease can be used for hydrolysis of fish meal, and the recombinant low-temperature neutral protease has higher enzymatic activity in the range of 15 DEG C to 35 DEG C, and still maintains more than 60% at 15 DEG C, so that the recombinant low-temperature neutral protease is a neutral low-temperature metal protease which can maintain higher activity at low temperature or normal temperature.

The invention relates to the technical field of microbial genetic engineering and fermentation engineering, and particularly discloses an aspergillus oryzae genetic engineering strain and application thereof, the aspergillus oryzae genetic engineering strain is ZW28-deltasA1 deltasB1 deltasC1 deltasD1, and is a quadruple gene knockout strain constructed by taking aspergillus oryzae ZW28 as an original strain and continuously knocking out protease genes AosedA, AosedB, AosedC and AosedD by using a gene editing technology. According to the quadruple gene knockout strain provided by the invention, the extracellular total protease activity is averagely reduced by 35%, the yields of expressed human lysozyme and calf chymosin respectively reach 65 mg / L and 122.2 mg / L, which are 4.7-5.2 times higher than those of an original strain, and the quadruple gene knockout strain shows strong universality for foreign proteins with different sources and pH characteristics, and provides a dominant platform for efficient production of industrial enzymes.