3310 results about "Mutant" patented technology

The invention belongs to the technical field of gene engineering and enzyme engineering, and particularly relates to an ornithine decarboxylase mutant as well as a coding gene, an expression vector, a recombinant strain and application thereof. The ornithine decarboxylase mutant is obtained by mutating wild ornithine decarboxylase ODC10 of which the amino acid sequence is shown as SEQ ID NO.1, the catalytic activity of the obtained mutant is improved, and the biosynthesis efficiency of 1, 4-butanediamine is favorably improved. The recombinant strain for expressing the ornithine decarboxylase mutant constructed by the invention can efficiently synthesize 1, 4-butanediamine, and the molar conversion rate of ornithine is remarkably improved, so that the production benefit is improved, and the production cost is reduced.

The invention belongs to the technical field of gene engineering, and particularly relates to mutants of murA and ygeA genes and application of the mutants in production of human milk oligosaccharides. According to the invention, murA and ygeA genes of escherichia coli are edited by using a gene editing technology, so that a genetic engineering chassis strain carrying murA and ygeA mutants is obtained. Further, the chassis strain is applied to the construction process of production strains of LNT II, LNnT, LNT, 3 '-SL and 6'-SL, and the yield of the LNT II, the LNnT, the LNT, the 3 '-SL and the 6'-SL is further increased.

The invention discloses a variation test method and device based on an intelligent automation script. The method comprises the following steps: extracting a statement execution sequence and a branch dependency relationship in an original source code by using a context-aware syntax tree, and generating a control flow diagram and a data flow diagram; performing code semantic analysis on the multi-dimensional code features by using a large language model, identifying potential defect types and variation rules, predicting the test efficiency of variants in combination with a deep Q learning model, and generating to-be-processed variants; performing grammar check and equivalence analysis on the to-be-processed variants to obtain effective variants, and testing the effective variants; variation test indexes are calculated, code weak points are positioned, and test blind areas are identified according to survival variants, so that a visual analysis report is generated; and iteratively generating a new test case according to the current survival variant and the visual analysis report, and testing the current survival variant by using the new test case. According to the invention, the variation test efficiency can be improved.

The present invention relates to the fields of genetic engineering and plant genetic breeding, and specifically to the application of wheat TaPND gene. In the present invention, by taking gene TaPND as a target, designing a sgRNA sequence based on CRISPR / Cas9, connecting a DNA fragment containing the encoding sgRNA sequence to a vector carrying CRISPR / Cas, a plant binary expression vector capable of knocking out the TaPND gene in wheat is constructed; by genetically transforming wheat, offspring with TaPND gene knockout are obtained, and wheat mutants with early flowering time are obtained. Experiments have confirmed that transgenic wheat with TaPND gene knockout achieves the advancement of wheat flowering time. The technical scheme of the present invention provides a practical method for realizing rapid wheat breeding using genetic engineering technology, which has important breeding application value and broad market application prospects.

The invention relates to peroxidase AniUPO or a mutant of the peroxidase AniUPO, application of the peroxidase AniUPO in synthesis of alpha-hydroxyl beta-ketoester derivatives and a method for synthesizing the alpha-hydroxyl beta-ketoester derivatives, and belongs to the field of enzyme engineering. The invention provides a peroxidase AniUPO mutant, which is an amino acid sequence obtained by taking wild type peroxidase AniUPO with an amino acid sequence as shown in SEQ ID NO.1 as a parent and mutating the wild type peroxidase AniUPO with the amino acid sequence as shown in SEQ ID NO.1. The invention also provides a DNA (Deoxyribonucleic Acid) molecule for coding the peroxidase AniUPO mutant, an expression vector, a host cell and a method for synthesizing the chiral alpha-hydroxyl beta-ketone ester derivative. Compared with a wild type, the peroxidase mutant has the advantages of higher catalytic activity and better selectivity, applicable substrate types can be expanded to various cyclic and non-cyclic beta-ketone ester substrates, expensive catalysts and ligands are not needed, reaction conditions are mild, and potential industrial application prospects are achieved.

The invention discloses application of a wheat WRKY transcription factor gene TaWRKY1, and belongs to the technical field of gene engineering. A nucleotide sequence of an open reading frame of the gene TaWRKY1 is shown as SEQ ID NO: 1, and the gene TaWRKY1 is infected by stripe rust to induce expression in interaction of wheat and stripe rust, and plays a negative regulation role in wheat stripe rust resistance. A TaWRKY1 gene editing mutant plant is created by using an agrobacterium tumefaciens-mediated genetic transformation method, the sporulation quantity on wheat leaves of the gene editing mutant is obviously reduced, and the allergic necrosis is increased, which indicates that the edited gene TaWRKY1 can improve the resistance of wheat to stripe rust. The invention provides a disease-resistant germplasm resource for the cultivation of the stripe rust-resistant wheat variety.

The invention discloses a glycosyl transferase UGT76G1 mutant and a method for catalytically synthesizing rebaudioside M by using the glycosyl transferase UGT76G1 mutant, and relates to the technical field of biological catalytic synthesis. The glycosyl transferase UGT76G1 mutant is any one of the following components (A)-(C): (A) a protein obtained by performing any one or more of the following mutations on the basis of an amino acid sequence as shown in SEQ ID NO.1: the 89th amino acid is mutated into H from M; the 380th amino acid is mutated from L to M; the 411th amino acid is mutated from A to Y; (B) a protein which has 95% or 98% or more of identity with the amino acid sequence defined by (A) and has the same function as the amino acid sequence defined by (A); and (C) a fusion protein obtained by connecting a tag to the terminal of the protein defined in (A) or (B). The glycosyl transferase UGT76G1 mutant disclosed by the invention is obtained by mutating and screening glycosyl transferase UGT76G1, has higher catalytic activity, and can be used for efficiently synthesizing rebaudioside M by taking rebaudioside D and UDPG as substrates.

The invention relates to the field of gene engineering, and discloses a D-3-phosphoglycerate dehydrogenase mutant, a coding gene thereof and application of the D-3-phosphoglycerate dehydrogenase mutant and the coding gene. Compared with a protein with an amino acid sequence as shown in SEQ ID NO: 1, the mutant comprises mutation at the 131 site and / or the 285 site. The mutant can specifically catalyze a substrate 3-phosphoglycerate to synthesize 3-phosphate-hydroxypyruvic acid, can effectively promote synthesis of L-serine in a metabolic pathway of microorganisms, improves the yield and conversion rate of the L-serine, and reduces the yield of byproducts at the same time.

The present invention discloses a Gs12-7 endonuclease variant and a gene editing system mediated thereby. Specifically, by means of a rational mutation strategy, two Gs12-7 mutants were constructed and compared, and it was found that after the 157th amino acid of the Gs12-7 endonuclease was mutated from Glu to Arg, the activity of this gene editing enzyme could be significantly improved, and this variant was designated as Gs12-7MAX. The present invention provides an efficient gene editing technology mediated by the CRISPR-Gs12-7MAX system, which has broad application prospects in the field of genome site-directed modification.

The invention discloses a glycosyl transferase UGTSL2 mutant. The UGTSL2 mutant is subjected to any one or more of the following mutations on the basis of an amino acid sequence as shown in SEQ ID NO.1: the 23rd amino acid is mutated from N to E; the 41st amino acid is mutated from R to P; the 91st amino acid is mutated from H to K; the 95th amino acid is mutated from K to D; the 123 amino acid is mutated from E to P; the 136th amino acid is mutated from L to F; the 151st amino acid is mutated from R to F; the 124 amino acid is mutated from H to E; the 168th amino acid is mutated from V to Y; the 198 amino acid is mutated from C to K; the 202nd amino acid is mutated from T to E; the 217th amino acid is mutated from W to K; the 225th amino acid is mutated from P to L; the 226 amino acid is mutated from F to V; the 285th amino acid is mutated from A to V; and the 333rd amino acid is mutated from I to V and the like.

The invention provides an UTR (Untranslated Region) element H2202 P1-G as well as a construction method and application thereof, and relates to the technical field of mRNA (messenger ribonucleic acid). According to the present invention, the ribosome load prediction and the secondary structure optimization are performed on the natural 5 'UTR of the HIV TAT 202 gene through the BaidleHelix platform, and the obtained HTAT 202 P1 sequence avoids the inhibitory hairpin structure so as to significantly improve the luciferase expression quantity compared to the natural UTR; an ncRNA sequence without a secondary structure is introduced on the basis of the HTAT 202 P1, translation inhibition of a 5 'cap region is further relieved, and the protein expression quantity of the constructed H2202 P1-G mutant (the DNA sequence of the H2202 P1-G is as shown in SEQ NO 1, and the RNA sequence is as shown in SEQ NO 2) is further improved.

The invention belongs to the technical field of enzyme engineering and biology, and particularly relates to a fungal luciferase mutant and application of the fungal luciferase mutant in improving the bioluminescence intensity of fungi or plants, and the amino acid sequence of the fungal luciferase mutant is shown as SEQ ID NO.1. The fungal luciferase mutant NnLuzT138P is obtained by mutating fungal luciferase NnLuz, the NnLuzT138P and other FBP luminescence pathway genes are co-expressed in fungi or plants, the bioluminescence intensity of the fungi and the plants can be remarkably improved, and compared with the prior art, the light quantum intensity is improved by more than two times. The invention provides a feasible scheme for construction of enhanced sustainable luminous fungi and creation of luminous plants.

The invention relates to the technical field of microbial genetic engineering, in particular to a sialyltransferase mutant with high yield of 3 '-sialyllactose as well as a construction method and application of the sialyltransferase mutant. The sialyltransferase mutant is selected from at least one of an NST protein or a mutant and a 406 NST protein or a mutant, the NST protein comprises an amino acid residue site in a sequence as shown in SEQ ID NO.1, and the 406 NST protein comprises an amino acid sequence in a sequence as shown in SEQ ID NO.2. The sialyltransferase mutant has the advantages that the sialyltransferase mutant can be used for preparing sialyltransferase, and the sialyltransferase mutant can be used for preparing sialyltransferase. The invention also provides a nucleic acid molecule containing the sialyltransferase mutant, an expression cassette, a recombinant vector, a recombinant microorganism and a construction method and application thereof, and has great significance in large-scale industrial production of 3-sialyllactose.

The invention belongs to the technical field of gene engineering and enzyme engineering, and particularly discloses an acid protease mutant and application thereof. According to the invention, genome DNA from Aspergillus niger is taken as a basis, a parent acid protease gene sequence is obtained through PCR amplification, random mutation is carried out through error-prone PCR, and an acid protease mutant with improved enzyme activity and a recombinant strain thereof are obtained through screening. After modification, under the optimal condition, the enzyme activity of the acid protease mutant is 251.64 U / mL, the enzyme activity is improved by 23.43% compared with that of parent acid protease (203.64 U / mL), meanwhile, the acid protease mutant is not prone to inactivation in the reaction process, has higher stability, can tolerate strong acid and high salt environments, shows a higher conversion rate in enzymolysis of collagen, and can be used for preparing the acid protease mutant. The leather softening agent has a good application prospect in leather softening.

The invention discloses a bovine I-type alpha interferon-ferritin fusion protein, a mutant thereof, a preparation method and an application of the bovine I-type alpha interferon-ferritin fusion protein. The bovine I-type alpha interferon is fused with a ferritin subunit, and interferon molecules are highly repeatedly and orderly displayed on the surface of a ferritin nanocage by utilizing the self-assembly characteristic of ferritin, so that the expression level, the structural stability and the antiviral activity of the interferon are remarkably improved. The fusion protein is further subjected to single-site or multi-site rational design mutation, and a mutant with significantly improved antiviral activity and stability is obtained. According to the invention, a silkworm or insect cell eukaryotic expression system is adopted to express the fusion protein or the mutant thereof, and the expression system is safe to operate, simple and convenient in procedure, low in cost and extremely beneficial to large-scale industrial production; the prepared fusion protein or mutant nanoparticles have application prospects in preparation of drugs or reagents for preventing or treating bovine viral diseases.

The invention discloses a Candida antarctica lipase B mutant with high transesterification activity and application thereof. The nucleotide sequence of the Candida antarctica lipase B mutant is shown as SEQ ID NO.1, and the amino acid sequence of the expression protein of the Candida antarctica lipase B mutant is shown as SEQ ID NO.2. The invention further discloses a preparation method of the Candida antarctica lipase B mutant. Compared with an original enzyme, the mutant has higher fat hydrolysis activity and transesterification activity, the specific enzyme activity of the fat hydrolysis activity reaches 480.39 U / mg, the specific enzyme activity of the transesterification activity reaches 40.67 U / mg, the biodiesel conversion rate at the temperature of 30 DEG C can reach 92%, low-temperature high catalytic activity is shown, and the mutant can be used for enzymatic preparation of biodiesel and has potential industrial application value.

The embodiment of the invention provides a tyrosine ammonia lyase mutant and application thereof in synthesis of p-coumaric acid. The mutant is subjected to any one of the following amino acid mutations on an amino acid sequence as shown in SEQ ID NO. 2: F497I, F497I + L248M, F497I + L248M + M4T, and L248M + F497I + M4T + V482L. According to the embodiment of the invention, an amino acid sequence as shown in SEQ ID NO. 2 is taken as a template, a key active site of the template is subjected to mutation transformation, a mutant library is constructed, and a series of mutants with remarkably improved catalytic activity and / or conversion rate are screened; the tyrosine ammonia lyase mutant provided by the embodiment of the invention can selectively catalyze L-tyrosine to generate p-coumaric acid, and has excellent yield and / or higher yield.

The invention provides a keto-alcohol acid reductoisomerase mutant and application of the keto-alcohol acid reductoisomerase mutant in construction of genetically engineered bacteria for high yield of D-pantothenic acid. In order to solve the technical problem that in the prior art, the yield of D-pantothenic acid synthesized through microbial fermentation is not high, semi-rational design of enzyme is adopted, and a keto-alcohol acid reductoisomerase mutant with substrate specificity is obtained and applied to construction of genetically engineered bacteria for high yield of D-pantothenic acid. According to the invention, a genetically engineered bacterium with high yield of D-pantothenic acid is constructed by utilizing the keto-alcohol acid reductoisomerase mutant through a CRISPR-Cas9 gene editing technology, so that the yield of D-pantothenic acid is increased by about 12.04%, and compared with an original strain, the valine accumulation amount in obtained fermentation liquor is almost unchanged compared with that of a control, and the yield of D-pantothenic acid is greatly improved. However, the accumulation of branched chain amino acid isoleucine is less, so that the modification of the substrate specificity of the keto-alcohol acid reductoisomerase is achieved.

The invention discloses an acid-resistant PET plastic degrading enzyme PETase mutant and application thereof, and belongs to the technical field of biological enzyme engineering. According to the invention, amino acid sequence truncation, single point mutation and pairwise compound mutation are carried out on the basis of wild type FAST-PETase, and the obtained mutant R84D reduces the optimal reaction pH of the FAST-PETase and improves the stability of the FAST-PETase under different pH values. Compared with the enzyme activity of starting FAST-PETase, the enzyme activity of the mutant constructed by the invention is improved by 2.2 times. When the pH value is 6.5-8.5, the stability of R84D is relatively high, and the enzyme activity residue is gt; 60%.

The invention discloses a hydroxysteroid dehydrogenase mutant with improved thermal stability, a screening method and application, key hotspot amino acid residues related to the thermal stability are rapidly identified based on computer-aided design technologies such as hydroxysteroid dehydrogenase structure prediction, PROSS and the like, so that a limited site-directed mutant library is constructed; and further screening to obtain a dominant mutant. The semi-inactivation temperature (T5105) of the obtained mutant is increased by 1.0-7.7 DEG C compared with that before mutation, wherein the semi-inactivation temperatures (T5105) of the mutants A126I and A132H are increased by 7.7 DEG C and 3.7 DEG C respectively. And the relative enzyme activity of the mutant reaches 99.86-136.73% of the relative enzyme activity before mutation. The invention provides a new method for rapidly improving the thermal stability of the enzyme, and lays a foundation for the industrial application of the hydroxysteroid dehydrogenase.

The invention belongs to the field of bioengineering, and particularly discloses a tryptophan synthetase mutant with improved enzymatic activity and application of the tryptophan synthetase mutant in production of L-cysteine. Compared with wild tryptophan synthetase from escherichia coli, the amino acid sequence of the tryptophan synthetase mutant has the advantages that a beta catalytic subunit has T69A, S143A, R219E and F280H site combined mutation. Researches show that the enzyme activity of the mutant is increased by 2.4 times compared with that of a wild type through enzyme activity determination and analysis of inducible expression and purification. The beneficial tryptophan synthetase mutant provided by the invention can efficiently biocatalyze L-serine and sodium hydrosulfide to synthesize L-cysteine, and has a better industrial application prospect.

The invention belongs to the technical field of enzyme engineering, and discloses an MHETase-EstD9 mutant and an application of the MHETase-EstD9 mutant. The invention provides an MHETase-EstD9 mutant, the amino acid sequence of the MHETase-EstD9 mutant is shown as SEQ ID No.2, and on the basis of the mutant, the amino acid of the MHETase-EstD9 mutant is further mutated to design five MHETase-EstD9 mutants. At 70 DEG C, compared with wild type MHETase-EstD9, the BHET degradation activity of the MHETase-EstD9 mutant provided by the invention is improved by 15% or more, and the highest BHET degradation activity can be improved by 24 times; meanwhile, the thermal stability of the MHETase-EstD9 mutant is also improved to a certain extent, and the Tm value is maximally improved by 18.0 DEG C. The method can be applied to the fields of degradation of PET, MHET or BHET, preparation of degradation agents of PET and degradation products thereof, TPA recovery and the like, and has important environmental benefits, economic benefits and social benefits.

The invention discloses a tryptophan synthase mutant as well as a coding gene, a vector, recombinant bacteria and application thereof. Comprising the following steps: carrying out error-prone PCR (Polymerase Chain Reaction) on a coding gene trpBA of TrpS from EscherichiaoliK12MG1655, screening mutants by combining high-throughput screening, and finding that the affinity of the TrpS to a substrate indole can be effectively improved by mutating 114-site Gln of a beta subunit of the TrpS into Met and mutating 84-site Gly into Ser, so that the synthesis of L-tryptophan is promoted. The efficiency of the TrpS for converting indole and L-serine into L-tryptophan is higher due to the improvement of the affinity of indole. A new idea is provided for efficient production of L-tryptophan in the fermentation industry.

The invention includes a mutant Taq polymerase, which can effectively amplify a target sequence under conditions of salt concentration(s) similar to body fluids, including blood, serum or plasma preserved with sodium citrate. The mutant Taq polymerase, or a biologically active fragment thereof, has one or more substitutions differing from the wild type as shown in Table I.

The embodiment of the invention provides a method for obtaining a tyrosine ammonialyase mutant based on high-throughput screening, the tyrosine ammonialyase mutant and application of the tyrosine ammonialyase mutant. The mutant is subjected to any one of the following amino acid mutations on an amino acid sequence as shown in SEQ ID NO.2: A245G + A247T, and D192V + A245G + A247T. According to the embodiment of the invention, an amino acid sequence as shown in SEQ ID NO.2 is taken as a template (basis), a key active site of the template is subjected to mutation transformation, a mutant library is constructed, and a series of mutants with remarkably improved catalytic activity and / or conversion rate are screened; the tyrosine ammonia lyase mutant provided by the embodiment of the invention can selectively catalyze L-tyrosine to generate p-coumaric acid, and has excellent yield and / or higher yield, which indicates that the mutant has more significant application prospects in catalysis of L-tyrosine to generate p-coumaric acid.

The invention discloses application of a soybean GmZG3 gene in regulation and control of soybean plant height, and belongs to the technical field of gene engineering. According to the present invention, Cas9-sgRNA and PTF-GmZG3 recombinant vectors of the soybean GmZG3 gene are constructed, the Cas9-sgRNA and PTF-GmZG3 recombinant vectors are transferred into agrobacterium to obtain EHA-GmZG3-sgRNA and EHA101 / PTF101-GmZG3 recombinant strains, and the constructed recombinant strains are converted into wild soybeans through an agrobacterium-mediated method so as to obtain the GmZG3 gene mutant and the overexpression plant. The phenotypes of the mutant and the overexpressed plant are identified, and the plant height of the mutant plant is obviously lower than that of the wild type soybean, and the plant height of the overexpressed plant is obviously higher than that of the wild type soybean. Therefore, the invention provides a method for genetically improving the soybean plant type, and the method has potential market value and wide application prospect in the agricultural field.

The invention relates to the technical field of biology, in particular to a leucine dehydrogenase mutant and application thereof. According to the leucine dehydrogenase mutant and the application thereof provided by the embodiment of the invention, wild leucine dehydrogenase is subjected to mutation screening to obtain the leucine dehydrogenase mutant with an amino acid sequence as shown in SEQ ID NO: 1 or SEQ ID NO: 2, the catalytic efficiency of asymmetric reductive amination of a prochiral alpha-keto acid compound is improved, and the thermal stability of the leucine dehydrogenase mutant is improved; the synthesis efficiency of the L-amino acid is favorably improved.

The invention discloses a haloalkane dehalogenase mutant and an application thereof in degradation of 1, 2, 3-tribromopropane, haloalkane dehalogenase DhaA31 coded by dhaA gene of a Rhodococcus rhodococcus NCIMB 13064 strain is modified to obtain the haloalkane dehalogenase mutant F168W / I246C, the haloalkane dehalogenase mutant has high degradation efficiency on haloalkane pollutant 1, 2, 3-tribromopropane, and the haloalkane dehalogenase mutant has high degradation efficiency on haloalkane pollutant 1, 2, 3-tribromopropane under the condition of water phase normal temperature. The degradation rate of the alkyl halide dehalogenase mutant to 10 mM of 1, 2, 3-tribromopropane within 1 h reaches 100%, and the degradation rate of the alkyl halide dehalogenase mutant to 10 mM of 1, 2, 3-tribromopropane reaches 100%. The haloalkane dehalogenase mutant has high stereoselectivity on (R)-2, 3-dibromo-1-propanol, and the e.r. Value of the haloalkane dehalogenase mutant reaches 6: 94. The haloalkane dehalogenase mutant provided by the invention has high catalytic efficiency and high stereoselectivity, is beneficial to recognition and further degradation of downstream enzymes, and has a good application prospect in bioremediation of emerging haloalkane pollutants.

The invention discloses a porcine epidemic diarrhea virus antigen fusion protein, a coding gene and a porcine epidemic diarrhea vaccine prepared from the porcine epidemic diarrhea virus antigen fusion protein. The method comprises the following steps: screening to obtain a homosensory sequence of the spike protein of the porcine epidemic diarrhea virus, and fusing the homosensory sequence with a monomeric porcine ferritin heavy chain subunit to obtain an antigen fusion protein; in order to improve the expression quantity or the titer of the antigen fusion protein, the obtained antigen fusion protein is subjected to sequence optimization and then subjected to single-site or multi-site mutation, and the expression quantity and the titer of the obtained mutant are remarkably improved. According to the invention, a recombinant protein is expressed by using a silkworm or AcMNPV-insect cell and other eukaryotic expression systems to obtain a self-assembled ferritin nanoparticle antigen for displaying an S protein antigen protein on the surface of a cage structure of porcine ferritin, and a vaccine prepared from the prepared nanoparticle antigen can induce a widely neutralized anti-porcine epidemic diarrhea virus antibody; the porcine epidemic diarrhea virus vaccine has the potential of becoming a universal porcine epidemic diarrhea virus vaccine with cross immune efficacy.

The invention discloses application of threonine dehydratase in regulation and control of growth of Didymeria segeticola and application of threonine dehydratase serving as a bactericide target in regulation and control of growth of Didymeria segeticola. The threonine dehydratase gene Td disclosed by the invention is derived from Didymeria segeticola, and a knockout mutant is obtained by constructing a knockout gene segment of the Td gene and introducing the knockout gene segment into a D.segeticola protoplast. Experiments find that the Td gene plays a role in growth of D.segeticola, in-vivo pyruvic acid content, in-vivo ATP (adenosine triphosphate) content and sensitivity to wuyiencin. Molecular docking analysis, molecular dynamics simulation and trace thermophoresis experiments prove that the threonine dehydratase can be used as a bactericide target, and the bactericide wuyiencin is obtained through screening. The threonine dehydratase disclosed by the invention can be used as a bactericide target and a key protein of a disease pathogenic mechanism, and has a wide application prospect in prevention and treatment of plant pathogenic fungal diseases.