5 results about "Point mutant" patented technology

This invention discloses a base editor, a base editing method, and its applications. First, the invention constructs highly efficient targeting sites unrestricted by PAM sequences through point mutation. A random insertion library is constructed using a transposase system, and proteins capable of cytosine base editing in mammals are screened using a BFP-GFP reporter system. Further point mutations are performed on deaminases to screen for highly efficient cytosine base editors unrestricted by PAM. The optimized base editing method of this invention achieves an efficiency of over 70%, with some sites reaching over 90%, demonstrating significant application value in basic life sciences, medicine, and other fields.

The application provides a formic acid dehydrogenase mutant, which is mutant I or mutant II; the mutant I is a single-point mutant based on wild-type formic acid dehydrogenase and is selected from one of R69H, K89E, R213H, R323H and K326E; the mutant II is a double-point mutant based on R69H and is selected from one of R69H / K89E, R69H / R213H, R69H / R323H and R69H / K326E. The application also provides a coding gene of the mutant and a recombinant expression vector and an engineering bacterium containing the gene. Meanwhile, the application also provides a preparation method of the mutant and an application of formic acid hydrogen production. The application obtains a mutant with high activity and substrate resistance by rational site-directed mutagenesis on wild-type formic acid dehydrogenase, significantly improves the catalytic activity and substrate resistance of the enzyme, improves the formic acid biological hydrogen production efficiency, reduces the production cost and is beneficial to industrial application.

The application provides a construction method and application of a CYP1A1 double-site mutant mouse model, and belongs to the technical field of genetic engineering. Specifically, the double-site mutant mouse model provided by the application is a CYP1A1 p.L118R and CYP1A1 p.R245P double-site mutant mouse model, which is obtained by the following method: S1: designing gRNA and a donor vector; S2: microinjecting Cas9 mRNA, gRNA and the donor vector into a fertilized egg; S3: obtaining F0 generation mice; S4: mating the positive F0 generation mice with wild-type mice to obtain F1 generation mice; S5: interbreeding the positive F1 generation mice to obtain F2 generation mice; and S6: screening the F2 generation mice to obtain the double-site mutant mouse model. The application can solve the technical problems of a complex preparation process and a low success rate of a double-site mutant mouse model far apart.

The present application relates to the technical field of genetic engineering and protein engineering, and particularly relates to a xylanase mutant. The mutant is a single-point mutant provided based on wild-type xylanase Xyn, and the specific activity of the mutant is generally increased by 12.4%-63.4%. The xylanase mutant containing the I128E single-point mutation has the highest specific activity of 1273.9 U / mg, which is conducive to reducing the production cost of the enzyme and promoting the wide application of the enzyme in the feed field.

This invention discloses a thermostable L-tryptophan oxidase mutant, belonging to the field of biotechnology. This invention analyzes information that improves enzyme thermostability from an evolutionary perspective, integrates and analyzes sequences of the thermostable reverse transcriptase family, and combines bioinformatics and crystallography methods to obtain a novel L-tryptophan oxidase mutant with high stability. The thermostable L-tryptophan oxidase mutants provided by this invention include single-point mutants and combined mutants. Compared with wild-type L-tryptophan oxidase, both single-point mutants and combined mutants have longer half-lives at 50°C; especially the combined mutants, which exhibit a synergistic effect of the thermostability of the single-point mutants, with a half-life approximately 8 times that of the wild type. The thermostable L-tryptophan oxidase mutants provided by this invention have excellent catalytic activity and promising application prospects.