35 results about "Insertional mutation" patented technology

The invention discloses enterobacter mori capable of resisting fish aeromonas and application of the enterobacter mori, the bacterial strain is classified and named as enterobacter mori SCH0241, the enterobacter mori SCH0241 is preserved in the China Center for Type Culture Collection on October 17, 2024, and the preservation number is CCTCC NO: M 20242236. The enterobacter mori SCH0241 disclosed by the invention is derived from the intestinal tract of high-stress-resistance healthy hybrid crucian carassius auratus No. 2, has good contact dependence antibacterial activity on aeromonas veronii, and can improve the capability of resisting aeromonas veronii infection of zebra fish. Through construction of an insertion mutation library and phenotype screening, it is found that a gene cluster of SCH0241, which exerts contact inhibition toxicity, is T6SS-2. The enterobacter mori SCH0241 disclosed by the invention provides a new powerful tool for biological prevention and treatment of fish diseases caused by aeromonas veronii.

The invention belongs to the technical field of microbial engineering, and discloses a method for producing eicosapentaenoic acid (EPA) with high yield by schizochytrium limacinum based on random mutation and low-temperature screening and application, and the method comprises the following steps: driving overexpression of a G418 resistance gene through a P2520 promoter, constructing a random insertion mutant library, and screening to obtain a schizochytrium limacinum mutant strain with a randomly mutated genome; coating in a culture medium containing G418 resistance for culturing, and screening out strains which are resistant to low temperature and good in growth state at 15 DEG C; and inoculating into a fermentation culture medium, adding MgSO4 with the final mass concentration of 0.1% at the initial stage of fermentation, supplementing MnCl2 with the final mass concentration of 0.05% when fermentation is performed for 72 hours, and culturing to obtain a mutant strain. According to the invention, the G418 resistance gene carried by the plasmid is randomly inserted into the schizochytrium limacinum genome, and low-temperature induced screening is combined, so that the target limitation of traditional homologous recombination is broken through, and a stable strain with multiple mutation superposition can be obtained through enrichment.

The present invention provides polynucleotide vectors for high expression of heterologous genes. Some vectors further comprise novel elements that further improve expression. The gene transfer systems can be used in methods, for example, gene expression, bioprocessing, gene therapy, insertional mutagenesis, or gene discovery.

Disclosed is a detection method for a dominant clone caused by an exogenous DNA insertion mutation, comprising: carrying out pretreatment, including plus-minus strand correction and redundancy integration, on virus IS information data; step 2, when the pretreatment is completed, identifying ISs caused by non-specific amplification so as to remove a false positive IS detection result caused by PCR non-specific amplification; step 3, merging the ISs at neighboring positions on the basis of a site clustering algorithm, and clustering the ISs; and step 4, analyzing a clone plane on the basis of the clustering result, and describing a dominant clone. According to the present invention, a PCR primer similarity ranking algorithm is utilized to identify and eliminate non-specific amplification potentially caused by PCR primer similarity, significantly reducing false positives of IS identification in practical applications, improving the accuracy.

The present disclosure provides methods of treating cancer in a patient determined to have an EGFR and / or HER2 exon 20 mutation, such as an insertion mutation, by administering a third-generation tyrosine kinase inhibitor, such as poziotinib or afatinib.

The application discloses a detection method based on dominant clones caused by exogenous DNA insertion mutation, and the method comprises the following steps: 1, preprocessing of virus IS information data by positive and negative strand correction and redundant integration; 2, after the preprocessing is completed, IS caused by non-specific amplification is identified to remove false positive IS detection results caused by PCR non-specific amplification; 3, according to a site clustering algorithm, IS in adjacent positions is combined and clustered; 4, according to the clustering results, a clone plane is analyzed, and dominant clones are described; and the application utilizes a PCR primer similarity ranking algorithm to identify and remove non-specific amplification caused by possible PCR primer similarity, so that the false positive of IS identification is significantly reduced in actual application, and the accuracy is improved.

The application relates to the technical field of gene editing, and provides a method for creating herbicide-resistant rice through gene editing of an intron region of an OsEPSPS gene, specifically, a WT / -22bp single allelic deletion mutation or a WT / -32+6bp single allelic deletion insertion mutation is introduced into the intron region of the OsEPSPS gene, that is, taking A in an initiation codon ATG of the OsEPSPS gene as the 0 position, 22bp is deleted from 771 to 792 downstream of the 0 position on one allele or 32bp is deleted from 775 to 806 downstream of the 0 position and 6bp is inserted into the interval, and the other allele remains unchanged. It is found by the application that the rice can obtain resistance to EPSPS inhibitor herbicides by introducing the above-mentioned mutation, the resistance is identified in T0 generation plants carrying the mutation, and can be stably inherited to the offspring plants, and the application value for improving the use efficiency of herbicides is extremely high.

The invention discloses an epegRNA (Ribonucleic Acid) primer pair for detecting an Indel marker related to porcine diarrhea virus resistance and application of the epegRNA primer pair. According to the epegRNA sequence provided by the invention, accurate AACA insertion mutation can be realized on a sequence of which the pig genome position is ChrX: 56448702 through pilot editing; the A / AAACA mutation can be used as a molecular marker to be applied to breeding for resisting porcine epidemic diarrhea.

The present application belongs to the technical field of fungal genetic breeding, and particularly relates to a molecular marker associated with the number of Lentinula edodes and application. The molecular marker is located at the 3865993th locus of chromosome 7 of the genome of Lentinula edodes, and is a 15bp insertion mutation. By using the molecular marker and detection method provided by the present application, the number of fruiting bodies (number of mushrooms) of Lentinula edodes at the mushroom stage can be efficiently determined at the mycelium stage, so as to accelerate the breeding process and improve the breeding efficiency. The molecular marker is a universal molecular marker, is less limited by genetic materials, and is widely applied.

The invention relates to the technical field of gene editing. Specifically, the invention provides a method for creating herbicide-resistant rice through gene editing of a 3 'UTR region of an OsEPSPS gene, and the method is characterized in that-38 / -13 + 12bp double allelic deletion insertion mutation is introduced into the 3' UTR region of the OsEPSPS gene, i.e., A in an initiation codon ATG of the OsEPSPS gene is taken as a 0 site, two alleles are respectively positioned at the 3363 to 3400 site deletion 38bp of the downstream of the OsEPSPS gene and at the 3377 to 3389 site deletion 13bp of the downstream of the OsEPSPS gene, and 12bp is inserted in the section. It is found that by introducing the double allelic deletion insertion mutation, rice can obtain resistance to EPSPS inhibitor herbicides, the resistance is identified in T0-generation plants carrying the mutation and can be stably inherited to offspring plants of the T0-generation plants, and the application value for guaranteeing agricultural production safety and improving the use efficiency of the herbicides is extremely high.

The invention discloses an arabidopsis thaliana VHA-B1 gene, a coding protein thereof, a mutant, and preparation, identification and application thereof, and the preparation of the mutant comprises the following steps: respectively targeting a ninth exon and / or a first exon of a VHA-B1 gene (such as SEQ ID NO.1) through a T-DNA insertion and / or gene editing technology to obtain an insertion mutant and / or a gene editing mutant, and carrying out amplification on the insertion mutant and / or the gene editing mutant to obtain the VHA-B1 gene. And identifying the insertion mutant and / or the gene editing mutant through PCR (Polymerase Chain Reaction) to obtain the homozygous mutant with VHA-B1 gene deletion. The tolerance of the obtained purified mutant to cucumber mosaic virus (CMV) is weakened, which shows that the Arabidopsis thaliana VHA-B1 gene can improve the CMV tolerance of the plant, and an application basis is provided for culturing antiviral crop varieties by using the VHA-B1 gene.

The present application discloses a strain of Enterobacter mori against Aeromonas veronii and its application, which is classified and named as Enterobacter mori (SCH0241). Enterobacter mori SCH0241, preserved in China Center for Type Culture Collection on October 17, 2024, with a preservation number of CCTCC NO: M 20242236. The Enterobacter mori SCH0241 of the present application is derived from the intestine of a highly stress-resistant healthy hybrid crucian carp “Hefang crucian carp No. 2”, has good contact-dependent bacteriostatic activity against Aeromonas veronii, and can improve the ability of zebrafish to resist Aeromonas veronii infection. Through the construction of an insertion mutation library and phenotype screening, it is found that the gene cluster of T6SS-2 of SCH0241 plays a contact inhibition toxicity. The Enterobacter mori SCH0241 of the present application provides a new powerful tool for the biological control of fish diseases caused by Aeromonas veronii.

To provide methods of treating cancer in a patient determined to have an EGFR and / or HER2 exon 20 mutation, such as an insertion mutation.SOLUTION: The present invention provides a method for treating cancer in a subject determined to have one or more EGFR exon 20 mutations, the method comprising a step of administering to the subject a third-generation tyrosine kinase inhibitor such as poziotinib or afatinib in an effective amount.SELECTED DRAWING: None

The invention relates to the technical field of gene editing, and provides a method for creating herbicide-resistant rice through gene editing of an intron region of an OsEPSPS gene, and particularly relates to a method for introducing single allelic deletion mutation of WT / 22bp or single allelic deletion insertion mutation of WT / 32 + 6bp into the intron region of the OsEPSPS gene, i.e., A in an initiation codon ATG of the OsEPSPS gene is 0 site, A in the initiation codon ATG of the OsEPSPS gene is 0 site, A in the initiation codon ATG of the OsEPSPS gene is 0 site, and A in the initiation codon ATG of the OsEPSPS gene is 0 site; on one allele, 22bp is deleted from the 771 site to the 792 site at the downstream of the allele or 32bp is deleted from the 775 site to the 806 site at the downstream of the allele, and 6bp is inserted in the interval, and the other allele is kept unchanged. It is found that by introducing the mutation, rice can obtain resistance to EPSPS inhibitor herbicides, the resistance is identified in T0-generation plants carrying the mutation and can be stably inherited to offspring plants of the T0-generation plants, and the application value for improving the use efficiency of the herbicides is extremely high.

The present disclosure provides compositions and methods of use of nucleic acids and complexes thereof targeted to nucleic acids. Genomic engineering can refer to the alteration of a genome by deletion, insertion, mutation, or replacement of a particular nucleic acid sequence. The alteration can be gene or position specific. Genomic engineering can utilize nucleases to cleave nucleic acids, thereby generating a site for alteration. Engineering of non-genomic nucleic acids is also contemplated.

The invention discloses an InDel marker located on a chromosome 9 of a pig and capable of regulating and controlling the papillary number heterosis of the pig and application of the InDel marker. The nucleotide sequence of the InDel marker is shown as SEQ ID NO: 1 or SEQ ID NO: 2; the site is located between the 79th site and the 80th site from the 5'end of the nucleotide sequence shown in SEQ ID NO: 1, and corresponds to insertion mutation of C between the 11197897 bp and the 11197898 bp on the No.9 chromosome of the international pig reference genome version 11.1. The InDel marker provided by the invention can be applied to crossbreeding of pigs so as to improve the nipple number heterosis of offspring pigs and realize genetic improvement of the pigs, and by breeding the pigs with the InDel marker genotype of T / T, the reproductive capacity of sows can be remarkably improved, and the number of weaned piglets can be increased.

The invention belongs to the technical field of biology, and discloses a plasmid for genetic manipulation of bacillus licheniformis and application of the plasmid. The temperature-sensitive plasmid pTSMK created by the invention can be transferred into bacillus licheniformis in a conjugational transfer manner, and has both high efficiency and universality. The plasmid is used for carrying target DNA (deoxyribonucleic acid), so that the genetic manipulation of the bacillus licheniformis can be realized, such as DNA knockout, DNA knockin and transposon random insertion mutation.

The invention relates to the technical field of plant genetic engineering application, and particularly provides application of a mutant transcription factor AHL14 gene in regulating and controlling the oil content of plant seeds and a method for improving the oil content of the plant seeds. According to the invention, the homozygous mutants ahl14-1 and ahl14-2 are obtained by identifying the DNA (deoxyribonucleic acid) and RNA (ribonucleic acid) levels of T-DNA insertion mutants of two AHL14 transcription factors. Phenotypic analysis shows that compared with wild Col-0, the total oil production of ahl14-1 and ahl14-2 mutant plant seeds is remarkably improved, and meanwhile, the seed size and thousand seed weight of the mutant plants are not reduced compared with those of the wild Col-0. The AHL14 gene disclosed by the invention provides a new excellent gene resource for molecular design breeding of oil crops, and has good reference value and application prospect in the aspect of high-oil breeding of the oil crops.

The invention discloses a circRNA-BMP2 delivery system for plant vesicles as well as a preparation method and application of the circRNA-BMP2 delivery system, and belongs to the technical field of biomedicine. According to the invention, the gastrodia elata vesicles are taken as a delivery carrier of circRNA-BMP2 and are fixed on the surface of the micro-arc oxidation implant, so that double-effect synergistic improvement of diabetes mellitus osseointegration is realized. Due to the beneficial components and the unique hollow biological membrane structure, the gastrodia elata vesicles are considered as potential candidate drugs for treating diseases, the plant-derived phospholipid structure of the gastrodia elata vesicles endows circRNA with a long-acting protection effect, and the risk of DNA insertion mutation is avoided. Meanwhile, the large-scale production cost of the gastrodia elata vesicles is reduced, and the gastrodia elata vesicles are free of animal The system breaks through the bottlenecks of short osteogenic factor release period, poor targeting property and unregulated inflammation microenvironment in the prior art, solves the problem that the inflammation microenvironment damages bone metabolism balance in a diabetic environment, and realizes long-acting slow release and stable expression of drugs.

The present invention discloses a phage genome editing vector based on the CRISPR-Cas9 system and its editing method and application. red9 A plasmid containing the gene and sgRNA elements is constructed, and specific primers are designed for the editing site to obtain the pTarget plasmid; the donor DNA sequence is constructed into a vector to obtain the pEdit plasmid. After phage is infected with a host bacteriophage containing the pEdit plasmid, the pTarget plasmid is used to reverse screen for mutant phage, achieving efficient and rapid gene editing of the phage genome, including gene deletion mutations, single nucleotide substitution mutations, and insertion mutations. This method is simple to operate and has high gene editing efficiency. It has promoted the development of structural analysis and functional research of Staphylococcus aureus phage genomes and provided a new strategy for the modification of engineered phages, with broad application prospects and market value.

The present invention relates to a new crystal form of Zipalertinib, a preparation method therefor, a pharmaceutical composition containing the crystal form, and the use of the crystal form in the preparation of an EGFR inhibitor drug and a drug for treating non-small cell lung cancer with an EGFR exon 20 insertion mutation.

The invention discloses arabidopsis thaliana myosin binding protein MyoBs and application thereof, and belongs to the technical field of biology. Mutant homozygous identification and screening are carried out on MyoB I (A) subfamily four members, pollen, pod length and seed phenotype of the homozygous inserted mutant are observed, it is found that flower morphology and pollen grain activity of a single mutant do not have abnormal phenotype, but the pod length of the single mutant is shorter than that of a wild type, and the single mutant has the advantages of being high in yield and high in yield. Therefore, the yield is also obviously lower than that of the wild type. According to the invention, phenotypes of short and small fruit pods and wizened seeds appear in a plant in an arabidopsis thaliana MyoBs protein single mutant, so that the importance of the MyoBs protein in a plant growth and development process is clarified, and a theoretical basis is provided for subsequent plant breeding.

The invention relates to the technical field of plant genetic engineering application, and particularly provides application of a mutant transcription factor AHL29 gene in regulating and controlling the oil content of plant seeds and a method for improving the oil content of the plant seeds. According to the invention, the homozygous mutants ahl29-2 and ahl29-3 are obtained by identifying the DNA (deoxyribonucleic acid) and RNA (ribonucleic acid) levels of T-DNA insertion mutants of two AHL29 transcription factors. Phenotypic analysis shows that compared with wild Col-0, the total oil production of ahl29-2 and ahl29-3 mutant plant seeds is remarkably improved, and meanwhile, the seed size and thousand seed weight of the mutant plants are not reduced compared with those of the wild Col-0. The AHL29 gene disclosed by the invention provides a new excellent gene resource for molecular design breeding of oil crops, and has good reference value and application prospect in the aspect of high-oil breeding of the oil crops.

The invention discloses a method for testing cross contamination between samples in a gene sequencing pretreatment platform, and relates to the technical field of molecular biological detection. The test method comprises the following steps: selecting a target gene with deletion mutation or insertion mutation; selecting a mutation site in the target gene; designing PCR (Polymerase Chain Reaction) amplification primers of the sample by utilizing base sequences before and after the mutation site; whether the negative sample has deletion mutation or insertion mutation is detected in a sequencing mode to judge whether the all-in-one machine has cross contamination prevention performance in the pretreatment process, if the frequency of deletion mutation or insertion mutation is larger than 0%, it is determined that cross contamination exists between the samples, and otherwise, it is determined that cross contamination does not exist between the samples. According to the test method, the problem of single base mutation caused by PCR amplification or sequencing errors does not exist, so that a false positive result is not generated, and an obtained judgment result is accurate; the method is simple in analysis process, clear in judgment standard and easy to implement.

This invention belongs to the field of biotechnology and discloses a plasmid for genetic manipulation of Bacillus licheniformis and its applications. The temperature-sensitive plasmid pTSMK created in this invention can be transferred into Bacillus licheniformis via conjugation transfer, exhibiting both high efficiency and versatility. Using this plasmid to carry target DNA, genetic manipulation of Bacillus licheniformis can be achieved, such as DNA knockout, DNA knock-in, and random transposon insertion mutations.

The invention belongs to the technical field of molecular biology and plant molecular breeding, and particularly relates to Camellia tonkinensis CdS-RNase gene insertion mutation and a molecular identification method and application thereof. On the basis that the earlier-stage CdS-RNase gene mutation can improve the selfing fruit setting rate of Vietnam camellia oleifera to a certain extent, the Vietnam camellia oleifera CdS-RNase gene 64bp insertion mutation is found for the first time, meanwhile, a primer pair is designed, a PCR amplification system and conditions are optimized, a rapid detection method for the CdS-RNase gene 64bp insertion mutation is provided, and an application example of the 64bp insertion mutation is provided. The screening efficiency of the Camellia tonkinensis CdS-RNase gene mutant can be improved, and the excavation process of the Camellia tonkinensis CdS-RNase gene mutant is accelerated.

The present invention refers to hyperactive variants of a transposase of the transposon system Sleeping Beauty (SB). The invention further refers to corresponding nucleic acids producing these variants, to a gene transfer system for stably introducing nucleic acid(s) into the DNA of a cell by using these hyperactive variants of a transposase of the transposon system Sleeping Beauty (SB) and to transposons used in the inventive gene transfer system, comprising a nucleic acid sequence with flanking repeats (IRs and / or RSDs). Furthermore, applications of these transposase variants, the transposon, or the gene transfer system are also disclosed such as gene therapy, insertional mutagenesis, gene discovery (including genome mapping), mobilization of genes, library screening, or functional analysis of genomes in vivo and in vitro. Finally, pharmaceutical compositions and kits are also encompassed.

The application discloses a Yarrowia lipolytica gene engineering bacterium for producing erythritol by glycerol, and the Yarrowia lipolytica gene engineering bacterium is obtained by random insertion mutation mediated by non-homologous end connection, and strains with high cell growth and high erythritol yield are screened. Further, the gene engineering bacterium overexpresses glycerol dehydrogenase gene GCY3 or ARA1 and dihydroxyacetone kinase gene DAK2 of a glycerol metabolism dihydroxyacetone pathway. Further, the gene engineering bacterium overexpresses transketolase gene TKL1 and transaldolase gene TAL1 of a non-oxidative pentose phosphate pathway. The application further discloses a construction method and application of the gene engineering bacterium. The superior gene engineering bacterium obtained by constructing a mutant library has the characteristics of tolerating high-concentration glycerol, and the substrate conversion rate and glycerol utilization rate are significantly higher than those of a starting strain.

The present invention relates to the use of ADAMTS1 gene polymorphisms for predicting the risk of Alzheimer's disease. Specifically, the present invention relates to the use of ADAMTS1 gene polymorphisms as markers for predicting the risk of Alzheimer's disease in a subject; preferably, the ADAMTS1 gene polymorphisms include a dupT insertion mutation at position 268474067415 of human chromosome 26 (i.e., rs3838078). On the other hand, the present invention relates to a kit for predicting the risk of Alzheimer's disease in a subject, the kit comprising one or more primers and / or probes for detecting ADAMTS1 gene polymorphisms.

The invention provides an FSH (follicle stimulating hormone) beta gene-based molecular marker for the reproductive performance of a Branchiotai Tibetan pig, a primer of the molecular marker and a method for screening a high-reproductive-performance Branchiotai Tibetan pig individual by using the marker, and the sequence of the molecular marker is a nucleotide sequence as shown in SEQ ID No: 1. The polymorphism of the insertion mutation site of the 292bp retrotransposon in the intron 1 of the FSH beta gene of the chang-dug Tibetan pig is shown as three genotypes, namely AA, AB and BB, wherein the AA type is a dominant genotype; a forward primer of a primer sequence of the molecular marker has a nucleotide sequence as shown in SEQ ID No: 2, and a reverse primer of the primer sequence of the molecular marker has a nucleotide sequence as shown in SEQ ID No: 3. The primer provided by the invention can specifically amplify the DNA sequence around the insertion mutation site of the 292bp retrotransposon in the intron 1 of the Tibetan pig FSH beta gene, the detection efficiency is high, and the reproductive performance of the Tibetan pig can be remarkably improved and the large-scale development of the Tibetan pig industry can be promoted when the primer pair is used for molecular marker-assisted breeding.