136 results about "Protease Gene" patented technology

The invention provides EV71virus-like particles as well as preparation method and application of the EV71virus-like particles. The method comprises the following steps of: connecting a P1 protein gene and 3CD protease gene of an EV71 virus with predicated mean vote (PMV) plasmid to build PMV-P1-3CD recombinant expression plasmid; turning into hansenula polymorpha AU-0501 expression bacterial strain to obtain an AU-PMV-P1-3CD recombinant expression bacterial strain; fermenting and culturing the recombinant expression bacteria strain and performing inducible expression on EV71 virus-like particle proteins by using methanol; centrifugally gathering thallus to perform homogenate crushing at high pressure; and purifying the supernate through ion-exchange column chromatography, hydrophobic chromatography and sieve chromatography to obtain the EV71virus-like particles. EV71virus-like particle vaccines provided by the invention has high immunogenicity, security, immunological characteristics and biological activity; and the EV71virus-like particles can be prepared in a large scale and purified and used for preparing vaccines for preventing EV71 infection, and has good economic value and application prospect.

The present invention relates to a method for the rapid and reliable detection of drug-selected mutations in the HIV protease gene allowing the simultaneous charaterization of a range of codons involved in drug resistance using specific sets of probes optimized to function together in a reverse-hybridization assay. More particularly, the present invention relates to a method for determining the susceptibility to antiviral drugs of HIV viruses in a biological sample, with said method comprising: a) if need be, releasing, isolating or concentrating the polynucleic acids present in the sample; b) if need be amplifying the relevant part of the protease gene of HIV with at least one suitable primer pair; c) hybrydizing the polynucleic acids of step a) or b) with at least one of the following probes: probes specifically hybridizing to a target sequence comprising codon 30; probes specifically hybridizing to a target sequence comprising codon 46 and / or 48; probes specifically hybridizing to a target sequence comprising codon 50; probes specifically hybridizing to a target sequence comprising codon 54; probes specifically hybridizing to a target sequence comprising codon 82 and / or 84; probes specifically hybridizing to a target sequence comprising codon 90; or the complement of said probes; further characterized in that said probes specifically hybridize to any of the target sequences presented in FIG. (1), or the complement of said target sequences; d) inferring from the result of step c) whether or not a mutation giving rise to drug resistance is present in any of said target sequences.

The invention provides a human serum glycated albumin assay kit which comprises a reagent 1 and a reagent 2, wherein the reagent 1 comprises the following components: 20-200mmol / L of buffer solution, 5-50KU / L of protease co-expression vector, 10-50KU / L of peroxisome, 2-20mmol / L of 4-amino antipyrine, 0.01-1% of preservative and 0.01-1% of stabilizer, and the protease co-expression vector is obtained by cloning a protease gene and an ascorbic acid oxidase gene onto a same vector for performing co-expression; and the reagent 2 comprises the following components: 20-200mmol / L of buffer solution, 5-50U / mL of fructose lysine enzyme, 1-10mmol / L of N, N-bis(4-sulfobutyl ether)-3-methylaniline, 0.01-1% of preservative and 0.01-1% of stabilizer. The kit can remove the interferences of globulin and ascorbic acid in human serum and has good stability and low cost.

The present invention relates to a method of producing a biologically active polypeptide having insulinotropic activity, the method comprising steps of: (a) transforming a genetically modified host cell that has protease gene knockout, with a polynucleotide vector encoding the polypeptide; and (b) growing the transformed host cell to produce the biologically active polypeptide; and a method of producing a biologically active polypeptide having an N-terminal recognition site His-Gly with insulinotropic activity, the method comprising steps of: (a) transforming a genetically modified Pichia pastoris that has protease gene STE13 knockout, with a polynucleotide vector encoding the polypeptide; and (b) growing the transformed Pichia pastoris to produce the biologically active polypeptide.

A recombinant gram-negative bacterial cell comprising one or more of the following mutated protease genes: a) a mutated Tsp gene, wherein the mutated Tsp gene encodes a Tsp protein having reduced protease activity or is a knockout mutated Tsp gene; b) a mutated ptr gene, wherein the mutated ptr gene encodes a Protease III protein having reduced protease activity or is a knockout mutated ptr gene; and c) a mutated DegP gene encoding a DegP protein having chaperone activity and reduced protease activity; wherein the cell is isogenic to a wild-type bacterial cell except for the mutated Tsp gene and / or mutated ptr gene and / or mutated DegP gene and optionally a polynucleotide sequence encoding a protein of interest.

There are provided hyperthermostable proteases having an amino acid sequences represented by SEQ ID NOs: 1, 3 and 5 of the Sequence Listing or functional equivalents thereof and hyperthermostable protease genes encoding those hyperthermostable protease. There is also disclosed a process for preparation of a hyperthermostable protease by culturing a transformant containing the gene.

The invention discloses a method for heterologous expression and purification of trichoderma harzianum acidic protease P6281. Trichoderma harzianum RNA is extracted and is subjected to reverse transcription to obtain cDNA; an acidic protease gene is obtained by PCR and is inserted into an expression vector; a recombinant plasmid is electro-transformed and introduced into pichia pastoris GS115 and induced expression is performed; and then a relatively pure enzyme product is obtained by a nickel column affinity chromatography and molecular sieve chromatography. The trichoderma harzianum acidic protease P6281 is successfully expressed and purified for the first time; the method improves the yield of the P6281 protease; and the enzyme activity of the obtained acidic protease is much higher than that of conventional trichoderma acidic protease.

A recombinant gram-negative bacterial cell comprising one or more of the following mutated protease genes: a) a mutated Tsp gene, wherein the mutated Tsp gene encodes a Tsp protein having reduced protease activity or is a knockout mutated Tsp gene; b) a mutated ptr gene, wherein the mutated ptr gene encodes a Protease III protein having reduced protease activity or is a knockout mutated ptr gene; and c) a mutated DegP gene encoding a DegP protein having chaperone activity and reduced protease activity; wherein the cell is isogenic to a wild-type bacterial cell except for the mutated Tsp gene and / or mutated ptr gene and / or mutated DegP gene and optionally a polynucleotide sequence encoding a protein of interest.

The invention belongs to the technical field of agricultural living things, and particularly relates to a fungal acid protease Bs2688 mutant and a gene and application thereof. An amino acid sequenceof the protease is as shown in SEQ ID No.3. The invention provides the novel protease gene, and the protease good in performance is produced by means of gene engineering, and can be applied to the industries of feed, food, medicines and the like.

The invention relates to a method for promoting plant growth. The method comprises the following steps: constructing a plant expression vector comprising FTO or other homologous demethylated protease gene; converting the plant expression vector into acceptor bacteria; converting the acceptor bacteria into target plants; culturing the obtained target plants, collecting and culturing the T0 generation seeds, screening positive plants from the seedlings; culturing the screened positive plants, after the growth of the seedlings, collecting the T1 generation seeds; culturing the T1 generation seeds, choosing positive plants from the seedlings; culturing the selected positive plants, after the growth of the seedlings, collecting the T2 generation seeds; culturing the T2 generation seeds, screening homozygous strain lines from the seedlings; and culturing the T1 generation seeds of the plants of the T2 generation homozygous strain lines so as to obtain the target plants. An exogenous gene that can modulate the RNA methylation modification of m6A in a cell is introduced into a plant through a transgene technology so as to modulate the m6A on the RNA of a plant in order to promote the plant growth.

The invention discloses a method for constructing a monascus strain capable of achieving high yield of acid protease. The method is characterized by firstly amplifying an Asp fragment of an acid protease gene in monascus from a monascus genome, then connecting the Asp fragment obtained through amplification to an empty vector to construct a recombinant expression vector of acid protease in monascus, transforming the recombinant expression vector of acid protease in monascus into agrobacterium tumefaciens to obtain recombinant agrobacterium tumefaciens and then guiding the recombinant agrobacterium tumefaciens into monascus by utilizing an agrobacterium tumefaciens-mediated method, thus obtaining the monascus strain capable of achieving high yield of acid protease. The monascus transformant strain capable of achieving high yield of acid protease, which is obtained by a gene recombination method, has the characteristics of high-efficiency expression, accuracy in processing and genetic stability of the transformant and can achieve passage stability under non-selective pressure. The expression quantity of the acid protease gene in the monascus strain capable of achieving high yield of acid protease is 3.30 times the expression quantities of wild type genes, so that high yield of acid protease can be achieved.