55 results about "Resistant mutants" patented technology

The invention belongs to the technical field of biological detection, and particularly discloses a targeted sequencing primer group and a kit for detecting common pathogens of children and drug-resistant genes thereof. The primer group comprises 145 primer pairs of common children pathogen primer groups and 67 primer pairs of drug-resistant gene primer groups of the common children pathogen primer groups. According to the kit provided by the invention, nucleic acid in a clinical sample is directly extracted, 212 pairs of specific primers are adopted to realize multi-targeted amplification of a pathogen target area, a high-throughput sequencing platform is adopted to accurately identify pathogens and drug-resistant mutant genes thereof, 72 common children pathogens and drug-resistant genes thereof can be detected at the same time, and the kit has a wide application prospect. More accurate diagnosis information is provided for clinic, a key technical support is provided for early-stage accurate treatment of children infectious diseases, and the technical problems that in the prior art, in children pathogen detection, the detection speed is low, the number of missed detection is large, and the drug-resistant blind area is large are solved.

The invention belongs to the field of pathogen detection, and particularly relates to a rapid detection kit for detecting mycoplasma pneumoniae and drug-resistant genes, the kit comprises: a mycoplasma pneumoniae upstream primer, a mycoplasma pneumoniae downstream primer, and a probe for detecting mycoplasma pneumoniae; the kit comprises an upstream primer and a downstream primer of the drug-resistant mutant gene at the 23S rRNA A2063G site of the mycoplasma pneumoniae, and a probe for detecting the drug-resistant mutant gene at the 23S rRNA A2063G site of the mycoplasma pneumoniae. The kit provided by the invention is shorter in actual amplification time and better in detection effect, can provide more accurate and more reference value detection results for clinicians, can complete detection under general instruments in the market, and is better in general applicability and wider in application range.

The invention relates to the technical field of molecular biology, in particular to a specific probe, a primer probe combination for detecting mycoplasma pneumoniae drug-resistant mutation and bordetella pertussis drug-resistant mutation and a kit. The primer probe combination and the kit provided by the invention can distinguish mutation at sites 2063 and 2064 of mycoplasma pneumoniae non-mutation sensitive strains and 23s rRNA genes as well as mutation at sites 2047 of bordetella pertussis non-mutation sensitive strains and 23s rRNA genes, and are relatively high in detection sensitivity; the sensitivity of a mycoplasma pneumoniae sensitive strain, a drug-resistant mutant strain and a bordetella pertussis sensitive strain can reach 25 copies / mL, high sensitivity and specificity are achieved by using locked nucleic acid modification at a specific position on a probe and specially optimized metal catalytic ions, and meanwhile, the reagent can be stably stored for 12 months at the temperature of 2-8 DEG C.

The invention belongs to the technical field of microbiological detection, and particularly relates to a real-time fluorescent quantitative PCR (polymerase chain reaction) primer, a probe and a kit for detecting a special drug-resistant pedigree of candida albicans. The nucleotide sequence of the specific primer F is as shown in SEQ ID NO. 1; the nucleotide sequence of the specific primer R is as shown in SEQ ID NO. 2; the nucleotide sequence of a probe matched with the primer for use is as shown in SEQ ID No.3. The kit prepared by the invention ensures high specificity and high sensitivity in a PCR amplification process, effectively distinguishes drug-resistant mutant strains from sensitive strains, keeps efficient detection capability in a low-copy-number DNA template, reduces non-specific amplification and background signal interference, and improves the reliability and accuracy of a detection result.

The invention discloses a locust microsporidia strain and a preparation and use method thereof, and relates to the technical field of locusts, the locust microsporidia strain and locust microsporidia have the preservation number of CGMCC No.19390 and are named as PL-GM1 strain. The preparation method of the locust microsporidia strain comprises the following steps: S1, screening high-temperature-resistant mutant strains; s2, other high-temperature-resistant microorganisms are used in a combined mode; s3, cultivating healthy insects; s4, healthy insect inoculation; s5, feeding diseases and pests; s6, microspores are processed and inspected, packaging and quality inspection are conducted, and finally freezing storage is conducted. According to the method, high-temperature-resistant mutant strains are screened, high-temperature-resistant culture can be conducted on the locust microsporidia of the PL-GM1 strain, and the locust microsporidia of the PL-GM1 strain can have the high-temperature-resistant characteristic; other high-temperature-resistant microorganisms are used in a combined mode, the microsporidia screened in the step S1 and the other high-temperature-resistant microorganisms are co-cultured, and the high-temperature-resistant capacity and the prevention and treatment effect of the microsporidia are enhanced.

The invention discloses a bamboo reed dormant bud radiation mutation breeding method which comprises the following steps: selecting robust bamboo reed stalks with high lignification degree and dormant buds, sawing the stalks into stalk sections section by section, performing 60Co-gamma ray radiation treatment on a part of the stalks, performing no radiation treatment on the rest of the stalks, and taking the stalks as a control group; putting the stalk sections with the dormant buds and the stalk sections with the dormant buds of the control group, which are subjected to radiation treatment, into an illumination culture room for rooting culture, performing water culture on the obtained seedlings in the illumination culture room, and performing cadmium-tolerant screening on the obtained water-cultured seedlings to obtain cadmium-tolerant seedlings. Observing and selecting survival transplanted seedlings with characters different from those of the control group in the whole growth process as primary radiation mutants; and carrying out asexual propagation on the obtained primary radiation mutant single plant, and carrying out secondary cadmium-resistant screening on the primary radiation mutant single plant to obtain the cadmium-resistant mutant. According to the method, the bamboo reed mutagenesis material can be subjected to early screening, the stably inherited mutant can be rapidly obtained, the operation is convenient, and the method has important significance on breeding of a new bamboo reed variety.

The invention belongs to the technical field of agricultural breeding, and particularly relates to a method for creating a malus spectabilis cold-resistant mutant by adopting an EMS mutagenic agent, which comprises the following steps: preparing peripheral calluses of malus spectabilis; performing EMS treatment on the peripheral calluses to obtain inoculated tissues; sequentially inoculating the inoculated tissue to an adventitious bud differentiation culture medium and a rooting culture medium for culturing to obtain a malus speciosa mutant; the method comprises the following steps: inoculating a malus prunifolia mutant to a hydroxyproline rooting seedling culture medium for culture, and then transferring the malus prunifolia mutant to a rooting seedling culture medium without hydroxyproline for recovery culture to obtain a hydroxyproline-resistant variant plant; and culturing into seedlings, and carrying out low-temperature stress and rewarming to obtain the malus speciosus cold-resistant mutant. According to the method, the treatment time and mutagenesis concentration of EMS mutagenesis are set, the half fatality rate and mutagenesis efficiency are guaranteed, the rooting seedlings obtained through EMS mutagenesis of the callus are continuously subjected to HYP directional screening of cold-resistant plants, the most suitable HYP selection pressure is obtained, the mutant screening efficiency is improved, and plants with higher cold resistance are obtained.

The invention relates to preparation and application of diarylamine compounds containing a propionamido indole structure. The compounds have a structural formula as shown in a general formula (I). The invention relates to a compound shown in a general formula (I), which has a strong effect of inhibiting L858R / T790M / C797S and Del19 / T790M / C797S drug-resistant mutant EGFR kinase, and also relates to application of the compound and pharmaceutically acceptable salt thereof in preparation of drugs for treating and / or preventing diseases caused by drug-resistant mutation of EGFR kinase, the invention particularly relates to application in preparation of medicines for treating and / or preventing cancers. The invention also provides a preparation method of the compounds, a pharmaceutical composition containing the compounds and application of the pharmaceutical composition. The compound is especially used for preparing drugs for treating and / or preventing lung cancer, and has good antitumor drug development and application prospects. (I)

The invention relates to diarylamine compounds containing a dimethyl phosphine oxide structure as well as a preparation method and application of the diarylamine compounds. The compounds have a structural formula as shown in a general formula (I). The invention relates to a compound shown in a general formula (I), which has a strong effect of inhibiting L858R / T790M / C797S drug-resistant mutant EGFR kinase, and also relates to application of the compound and pharmaceutically acceptable salt thereof in preparation of drugs for treating and / or preventing diseases caused by drug-resistant mutation of EGFR kinase, the invention particularly relates to application in preparation of medicines for treating and / or preventing cancers. The invention also provides a preparation method of the compounds, a pharmaceutical composition containing the compounds and application of the pharmaceutical composition. The compound is especially used for preparing drugs for treating and / or preventing lung cancer, and has good antitumor drug development and application prospects.

The invention relates to a corn WRKY transcription factor ZmWRKY74 as well as a coding gene and application thereof, and belongs to the technical field of plant gene breeding, the CDS sequence of the coding gene of the transcription factor ZmWRKY74 is as shown in SEQ ID NO.1, the gene negatively regulates the drought tolerance of crop plants, and the amino acid sequence of the transcription factor ZmWRKY74 is as shown in SEQ ID NO.3. Gene positioning analysis is carried out on a drought-resistant mutant of corn, it is found that the ZmWRKY74 gene is an effect gene capable of improving the tolerance of corn to drought stress, and through a ZmWRKY74 gene overexpression experiment, the negative regulation effect of the ZmWRKY74 gene on the drought tolerance of corn plants is verified; important theoretical significance and application value are provided for creation of new corn germplasm and molecular breeding of corn.

The invention discloses a novel application of a pyrrolone benzenesulfonamide compound, the compound has obvious antiviral activity and low cytotoxicity, can be used as an allosteric inhibitor of coronavirus 3CL protease (3CLpro), and especially shows efficient inhibition capability on SARS-CoV-2 3CLpro. An enzyme activity test and molecular dynamics simulation result shows that the compound is combined with a 3CLpro allosteric site to induce the volume of a catalytic pocket to shrink, so that enzyme activity inhibition is realized, and the drug resistance risk caused by active site mutation is avoided. In-vitro antiviral experiments further show that the compound has a strong inhibition effect on SARS-CoV-2 wild type and various drug-resistant mutant strains (such as E166V and S144M), the EC50 value is as low as the nanomole level, the selection index is high, and the safety is good. Besides, based on the high conservative property of 3CLpro in different coronaviruses, the compound provided by the invention also shows inhibitory activity on other coronaviruses 3CLpro such as SARS-CoV, MERS-CoV and the like, and has broad-spectrum antiviral potential. The compound can be used for preparing medicines for preventing, relieving or treating related diseases caused by coronavirus infection, and has a good clinical application prospect.

The invention belongs to the technical field of bamboo reed breeding, and discloses a mutation breeding method for obtaining a bamboo reed cold-resistant mutant based on EMS (Enhanced Mass Spectrometry), which comprises the following steps: step 1, inducing embryogenic callus; step 2, carrying out EMS mutagenesis treatment; step 3, performing EMS stress screening; 4, differentiation and rooting culture of adventitious buds; 5, screening the tissue culture seedlings at low temperature; the mutation obtained through mutagenesis treatment is a permanent change of a genetic structure of an organism and may appear in germ cells and somatic cells of a plant. Physical or chemical mutagenic agents are used to increase the frequency of mutations. In chemical mutagens, EMS is considered to be the most effective mutagen, the EMS can generate a large number of non-lethal point mutations in the whole genome, the mutation frequency is high, the chromosome abnormality frequency is low, the EMS is used for treating embryogenic calluses of the bamboo reeds, Hyp is used for screening, the low-temperature-resistant bamboo reed mutants are obtained, the planting area of the bamboo reeds can be enlarged, and the survival rate of the bamboo reeds is increased. And waste land in a low-temperature region is fully utilized.

The application belongs to the field of genetic engineering and medical engineering, and discloses virK Application of gene in regulation of Klebsiella pneumoniae virulence. The application finds that the expression level of the gene is significantly increased in polymyxin-resistant mutant strains through transcriptome analysis, virK Further research shows that the expression of the gene is regulated by the PhoP / PhoQ two-component regulatory system. The application constructs virK Gene deletion mutant strains and complementary strains, and compares the virulence difference of the strains in an animal infection model, finds that virK The virulence of the deletion strain is significantly reduced, and the complementary strain can restore the virulence phenotype, indicating that virK The gene plays an important role in the regulation of Klebsiella pneumoniae virulence. The application first finds that PhoP can regulate virK Gene expression, and affects the virulence phenotype of Klebsiella pneumoniae through the regulation pathway, thereby revealing a new virulence regulation mechanism, and providing a theoretical basis for subsequent targeted intervention.

The present invention provides arylphosphine oxides with kinase inhibitory activity, which can effectively inhibit the activities of various types of EGFR resistant mutants (such as EGFR del19 , EGFR del19 / T790M , EGFR del19 / C797S , EGFR T790M / L858R , EGFR L858R / C797S , EGFR del19 / T790M / C797S , EGFR L858R / T790M / C797S ), and also has a significant inhibitory effect on the ALK fusion gene and mutants (such as L1196M), and can be used for the treatment, combination therapy or prevention of various cancers.

The invention discloses an acid-resistant cyclodextrin glycosyl transferase (CGTS) mutant and an application of the acid-resistant cyclodextrin glycosyl transferase CGTase mutant. The acid-resistant cyclodextrin glycosyl transferase CGTase mutant has the advantages that the acid-resistant cyclodextrin glycosyl transferase CGTase mutant can be used as an acid-resistant mutant; the CGTase is subjected to multi-site directed mutation based on an artificially designed wild type sequence (the amino acid sequence is as shown in SEQ ID NO: 2), and an acid-resistant mutant (the amino acid sequence is as shown in SEQ ID NO: 16) with the optimal reaction pH of 5.0 is obtained. The mutant can maintain more than 80% of catalytic activity in a pH range of 4.0-5.5, and can be stabilized for 10 hours under a strong acid condition (pH is 4.0). Compared with a wild type, the mutant has a higher conversion rate, a shorter reaction period and better substrate stability in the process of synthesizing 2-O-alpha-D-glucopyranosyl-L-ascorbic acid (AA-2G), the industrial production efficiency of the AA-2G can be remarkably improved, and the mutant has an important application value.

The invention relates to the technical field of herbicide resistance, in particular to a method for screening herbicide-resistant rice by using molecular markers, which comprises the following steps: S1, screening herbicide-resistant mutants, and extracting genome DNA of rice seedling leaves by using a CTAB (Cetyltrimethyl Ammonium Bromide) method; s2, specific primers are designed for ALS gene mutation sites, and flanking sequences of the mutation sites are covered. And S3, amplifying the target fragment by using PrimeSTAR Max enzyme, and verifying the product through agarose gel electrophoresis. And S4, carrying out DNA sequencing on the PCR product, and comparing ALS gene sequences to detect SNP locus mutation. And S5, according to the existing combination of the mutation sites Gln547Leu, Glu549Lys and Glu604Asp, judging the resistance of the plant to bispyribac-sodium and quinclorac. And S6, spraying mixed herbicides bispyribac-sodium and quinclorac to the mutant, and confirming the resistance through the survival rate and phenotype. The method can be directly adapted to a rice direct seeding cultivation system for non-transgenic resistant rice varieties, the weed competition pressure is remarkably reduced through directional weeding in the seedling stage, and an innovative solution is provided for solving the problem of weed damage in the direct seeding rice field.

The invention provides a methanol-tolerant Eimenococcus sp. Mutant strain as well as a screening method and application thereof, the methanol-tolerant Eimenococcus sp. Mutant strain is preserved in the China Center for Type Culture Collection (CCTCC), and the preservation number is CCTCC M 20232483. The mutant strain can effectively utilize an industrial byproduct methanol as a carbon source to carry out efficient protein production and valuable carotenoid accumulation; the mutant strain shows excellent photosynthesis capability and nitrogen assimilation efficiency under a methanol culture condition. Effective technical support is provided for methanol-based industrial biological refining application of microalgae, and the method has remarkable environmental and economic benefits.

The application discloses application of SGI-7079 in treating FLT3 mutant acute myeloid leukemia. The application research shows that SGI-7079 can be stably combined with FLT3, inhibit the phosphorylation level of FLT3 and downstream STAT5, AKT and ERK, and induce G1 phase arrest and cell apoptosis. In vitro, SGI-7079 has strong inhibitory effect on FLT3-ITD and drug-resistant mutant cells; in a FLT3-ITD mouse model, SGI-7079 can significantly reduce leukemia load and prolong survival, and shows better curative effect than existing drugs on drug-resistant mutations such as F691L and D835Y. Meanwhile, SGI-7079 also has significant inhibitory effect on primary cells of FLT3-ITD positive patients. It is shown that SGI-7079 can be used as a new drug candidate for treating FLT3 mutation and drug-resistant AML, and provides a new direction for optimizing targeted therapy strategy.

The invention provides a temperature-resistant breeding method for a temperature-resistant strain capable of synchronously removing sulfur and nitrogen. The method comprises the following steps: performing susceptible state culture on the strain capable of removing sulfur and nitrogen to change the selectivity and permeability of bacterial cells to a mutagenic agent; then, diethyl sulfate is added, more diethyl sulfate is promoted to enter cells under the action of alternate ice bath and 45 DEG C heat shock, and mutagenesis is carried out on the sulfur and nitrogen removal strains; and inoculating the mutated bacterial liquid into an inorganic culture medium salt solution containing Mg < 2 + >, and repeatedly culturing in a dark environment at 45 DEG C to obtain the temperature-resistant mutant strain with fixed temperature-resistant mutant inheritable characters. The mutation rate of the strain obtained by the breeding method is greatly improved and can reach 24.5% at most, the sulfur removal rate at 45 DEG C can reach 87.8%, and the strain has excellent temperature tolerance and sulfur and nitrogen removal rate.

The present invention discloses a substituted 2-arylaminopyrimidine compound, a pharmaceutical composition, and its use, belonging to the field of chemical medicine. The present invention provides a novel 2-arylaminopyrimidine compound having a structure represented by general formula (I). This compound has enhanced metabolic stability and a longer metabolic half-life, exhibits higher inhibitory activity against activated or drug-resistant mutant forms of EGFR than wild-type EGFR, and can effectively reduce side effects. #imgabs0#

The present invention relates to certain 2-(2,4,5-substituted-anilino)pyrimidine compounds and pharmaceutically acceptable salts thereof which may be useful in the treatment or prevention of a disease or medical condition mediated through certain mutated forms of epidermal growth factor receptor (for example the L858R activating mutant, the Exon19 deletion activating mutant and the T790M resistance mutant). Such compounds and salts thereof may be useful in the treatment or prevention of a number of different cancers. The invention also relates to pharmaceutical compositions comprising said compounds and salts thereof, especially useful polymorphic forms of these compounds and salts, intermediates useful in the manufacture of said compounds and to methods of treatment of diseases mediated by various different forms of EGFR using said compounds and salts thereof.

This invention belongs to the field of rice molecular breeding technology and discloses genes regulating cold and salt tolerance in rice seedlings. SOP17 Its applications. This gene has the nucleotide sequence shown in SEQ ID NO.1, and the amino acid sequence of the protein it encodes is shown in SEQ ID NO.2. This invention obtained a seedling-stage cold-resistant mutant of indica rice 9311 through EMS mutagenesis. sop17-1 The clone was obtained by analyzing the Mutmap. SOP17 Genes. Constructed using CRISPR / Cas9 gene editing technology. SOP17 Gene knockout vector, to obtain SOP17 Homozygous mutant lines. Functional validation showed that... SOP17 SOP17 This gene simultaneously negatively regulates cold and salt tolerance in rice seedlings. Knocking out this gene significantly enhances the tolerance of rice seedlings to low temperature (8°C) and salt (150mM NaCl). This invention provides new gene resources and molecular targets for breeding stress-resistant rice varieties.

The invention belongs to the technical field of vaccine preparation, and particularly relates to an attenuated vaccine resistant to tomato chlorosis virus and tomato yellow leaf curl virus as well as a construction method and application of the attenuated vaccine. The attenuated vaccine comprises a tomato chlorosis virus resistant mutant and a tomato yellow leaf curl virus resistant mutant, the mutant resistant to the tomato chlorotic virus comprises a gene segment which is any one of gene segments as shown in SEQ ID NO.1-5. The mutant resistant to the tomato chlorotic virus is a mutant resistant to the tomato chlorotic virus. The mutant resistant to the tomato yellow leaf curl virus comprises a gene segment as shown in SEQ ID NO. 6 or SEQ ID NO. 7. The vaccine prepared according to the method disclosed by the invention not only can resist skeleton vector viruses, but also can resist tomato chlorosis viruses and tomato yellow leaf curl viruses, and provides technical support for prevention and treatment of single infection and composite infection of ToCV and TYLCV.

The invention discloses a heat-resistant α-L-rhamnosidase and its sugar-tolerant mutant and application, belonging to the technical field of genetic engineering and enzyme engineering. The invention clones a heat-resistant α-L-rhamnosidase gene from the genome of Dictyoglomus thermophilum DSM 3960, and the nucleotide sequence is shown in SEQ ID NO.1. Compared with the prior art, the α-L-rhamnosidase DthRha provided by the invention has excellent heat resistance, and the heat-resistant α-L-rhamnosidase DthRha is subjected to sugar-tolerant key site mutation, and the obtained novel α-L-rhamnosidase DthRha sugar-tolerant mutant has a sugar tolerance coefficient increased by more than 2.3 times, and the mutant can specifically hydrolyze rhamnose connected by α-1 glycosidic bond at the R2 position of icariin, and can utilize the mutant to efficiently transform icariin to generate icariin I.

The invention provides a clostridium beijerinckii oxygen-resistant mutant strain as well as a construction method and application thereof. The clostridium beijerinckii mutant strain provided by the invention is a mutant obtained by inactivating a PerR protein coding gene Cbei1336 due to fixed-point insertion. The mutant strain not only can grow under an anaerobic condition, but also can well grow in aerobic air, is oxygen-resistant, has remarkably improved performances of producing butanol and acetone, and shortens the fermentation period by 24 hours.

The invention provides a pharmaceutical composition for degrading a PML-RAR alpha protein drug-resistant mutant and application of the pharmaceutical composition. The pharmaceutical composition comprises interferon and arsenic trioxide. According to the composition disclosed by the invention, HeLa cells or NB4 cells for expressing the drug-resistant mutant of the PML-RAR alpha fusion protein are jointly treated by using interferon and arsenic trioxide, so that the degradation of the drug-resistant mutant of the PML-RAR alpha fusion protein is specifically induced. The invention provides help for researching the degradation mechanism of the PML-RAR alpha fusion protein drug-resistant mutant and developing drugs for patients with arsenic drug-resistant APL.

This invention relates to the fields of enzyme engineering and bioengineering, and particularly to the bioengineering technology of an MHET hydrolase. This invention discloses a mutant MHET hydrolase, wherein the amino acid at position 66 of IsMHETase is mutated from Ala to Pro, amino acid at position 136 from Ser to Glu, amino acid at position 159 from Thr to Val, amino acid at position 192 from Met to Phe, amino acid at position 330 from Ala to Lys, amino acid at position 333 from Ala to Arg, amino acid at position 328 from Thr to Arg, amino acid at position 410 from Gln to Phe, amino acid at position 413 from Ser to Asn, amino acid at position 434 from Asp to Thr, amino acid at position 501 from Ala to Ile, amino acid at position 509 from Ala to Lys, and amino acid at position 561 from Ser to Val; the amino acid composition of the MHET hydrolase IsMHETase is shown in SEQ ID No. 1 of the sequence listing. Compared with wild-type MHET hydrolase, the heat-resistant mutant provided by this invention can be used in combination with PET hydrolase at higher temperatures, and its PET degradation activity is significantly improved.

This invention discloses a method and its application for creating stress-resistant mutants of common bean using chemical mutagenesis. The main steps include: determining the screening concentration of hydroxyproline (Hyp), determining the median lethal dose of ethyl methanesulfonate (EMS), and subculturing the stress-resistant mutants. After 3-5 subcultures, the stress-resistant mutants screened by this invention ultimately yielded 31 cotyledonary nodes of common bean resistant to hydroxyproline (Hyp), with a mutation rate of 8.16%. This invention can induce mutagenesis and directionally screen for stress-resistant common bean mutants, providing material support for stress-resistant breeding of common bean.

The invention provides a rice OsACC herbicide-resistant mutant gene double-marker rapid detection method and a kit, and belongs to the technical field of resistance analysis. The invention provides a double-labeled primer group which is used for detecting 11672nd base mutation in a coding sequence of a 34th exon CT structural domain (1653bp) of a rice OsACC gene, the mutation can endow a plant with ACCase inhibitor herbicide resistance, the invention specifically discloses a targeting fragment on the gene, a wild type fragment comprises a sequence as shown in SEQ ID No.1, the wild type fragment comprises a sequence as shown in SEQ ID No.2, and the wild type fragment comprises a sequence as shown in SEQ ID No.3 and a sequence as shown in SEQ ID No.4. The mutant fragment comprises a sequence as shown in SEQ ID No. 2. The invention constructs a dual molecular marker detection system, can solve the problem of low resolution or strong equipment dependence of single molecular marker type detection, has the advantages of high throughput, low cost and high accuracy, and provides a basis for herbicide resistance identification and precise herbicide selection prevention and control of resistant weeds.

The invention discloses a cutinase thermal stability mutant based on directed evolution and computer-aided design transformation and application thereof, and belongs to the technical field of enzyme engineering. Through iterative recombination of the system, the heat-resistant mutant M8 with the catalytic activity maintained is obtained through screening. Compared with a wild type, the half-life periods of the mutant M8 at 65 DEG C and 70 DEG C are respectively improved by 8110.4 times and 3982.0 times, and the mutant M8 shows excellent stability enhancement. The mutant M8 constructed on the basis respectively keeps 76.85% and 68.15% of activity under the alkaline conditions of pH 10.5 and 11, the thermal stability and the stability in an alkaline environment are remarkably enhanced, and the application potential of the mutant M8 under the high-temperature and alkaline conditions is widened.