1266 results about "Genome editing" patented technology

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.

Provided are compositions for genome editing and site-directed integration in plants comprising microprojectile particles coated, treated of applied with a recombinant DNA construct comprising a sequence encoding one or more genome editing reagents for delivery to a mature embryo explant from dry seeds. Further provided are methods for genome editing and site-directed integration in at least one cell of a plant using the disclosed compositions, and plants, plant parts and seeds comprising an edited genome or site-directed integration, which are produced by the disclosed methods.

The specification provides methods for introducing a desired genetic change in a nucleotide sequence using a double-strand break (DSB)-inducing genome editing system, the method comprising: identifying one or more available cut sites in a nucleotide sequence; analyzing the nucleotide sequence and available cut sites with a computational model to identify the optimal cut site for introducing the desired genetic change into the nucleotide sequence; and contacting the nucleotide sequence with a DSB-inducing genome editing system, thereby introducing the desired genetic change in the nucleotide sequence at the cut site.

The present disclosure is directed to controlling pest infestation by inhibiting one or more biological functions in an invertebrate pest. The disclosure discloses methods and compositions for use in controlling pest infestation by feeding one or more different recombinant double stranded RNA molecules to the pest in order to achieve a reduction in pest infestation through suppression of gene expression. The disclosure also discloses methods and compositions for targeted genome editing in the pest in order to achieve a reduction in pest infestation through disruption of protein activity. The disclosure is also directed to methods for making transgenic plants that express the double stranded RNA molecules and targeted genome editing constructs for use in protecting plants from pest infestation.

The invention discloses an application of a soybean nucleolus GTP binding protein gene GmNSN1 in soybean resistance to soybean mosaic virus diseases. The invention relates to an application of a soybean nucleolus GTP binding protein gene GmNSN1 as shown in SEQ ID NO.1 and SEQ ID NO.2 in genetic engineering modification of soybean mosaic virus resistance. The soybean nucleolus GTP binding protein gene GmNSN1 can negatively regulate and control the resistance of soybeans to diseased soybean mosaic virus diseases, and the antiviral response of the soybeans can be activated due to function deficiency of the soybean nucleolus GTP binding protein gene GmNSN1. The gene is knocked out through a CRISPR / Cas9 gene editing technology, so that the resistance of soybeans to the soybean mosaic virus disease can be remarkably improved.

The invention discloses a toxoplasma gondii attenuated vaccine strain RH delta rop67 as well as a construction method and application thereof, and belongs to the technical field of parasitic disease prevention and control and biological product preparation. The attenuated vaccine strain is constructed by performing targeted knockout on the ROP67 gene in a toxoplasma gondii strain RH delta ku80 through a CRISPR / Cas9 mediated gene editing technology. Compared with a wild type strain, the attenuated vaccine strain shows remarkable attenuation characteristic and good immunogenicity. A test result shows that the attenuated vaccine strain can induce a host to generate specific immune response mainly based on cellular immunity, maintains a protection effect on toxoplasma gondii infection in a relatively long immune period, and has a protection effect on tachyzoite infection and a chronic infection stage of toxoplasma gondii strains with different virulence; the survival ability of a host to tachyzoite infection can be improved, and the formation level of cysts in tissues is reduced. The invention provides a technical scheme with long-term immune potential for research and development of toxoplasma gondii attenuated vaccines.

The invention discloses a rice salt stress resistant gene OsHAK11 coding protein and application thereof, and belongs to the field of plant genetic engineering. According to the method, a rice OsHAK11 gene (the nucleotide sequence is shown as SEQ ID NO.1) is knocked out through a CRISPR / Cas9 gene editing technology, and a mutant with significantly enhanced salt tolerance is obtained. The gene editing vector pEGCas9Pubi-B-OsHAK11 contains sgRNA of a region as shown in a target SEQ ID NO.4, a rice receptor material is transformed through agrobacterium tumefaciens mediation, and a plant with the OsHAK11 gene subjected to frame shift mutation is obtained through screening. A salt stress experiment shows that the survival rate of the mutant oshak11 is obviously higher than that of a wild type. The invention provides a new gene resource and an efficient technical means for salt-tolerant breeding of rice.

The invention discloses an application of a soybean ARGONAUTE family gene GmAGO5 in soybean resistance to soybean mosaic virus diseases. The invention also discloses application of the soybean ARGONAUTE family gene GmAGO5 as shown in SEQ ID NO.1 and SEQ ID NO.2 in genetic engineering modification of soybean mosaic virus resistance. The soybean ARGONAUTE family gene GmAGO5 can negatively regulate and control the resistance of soybeans to diseased soybean mosaic virus diseases, and the antiviral response of the soybeans can be activated due to function deficiency of the soybean ARGONAUTE family gene GmAGO5. The gene is knocked out through a CRISPR / Cas9 gene editing technology, so that the resistance of soybeans to the soybean mosaic virus disease can be remarkably improved.

The invention belongs to the technical field of gene editing, and discloses a gene editing system and application. The invention provides a gene editing system. The gene editing system comprises Cas9 nickase, sgRNA (small guide ribonucleic acid) and annular petRNA. According to the present invention, the target chain cutting mediated reverse pilot editing (Split Reverse Prime Editing, srPE) realizes the precise editing of the non-target chain cutting site upstream sequence, such that the gene editing coverage range is expanded from the specific region only limited to the non-target chain cutting site downstream to the whole genome range, the stable and efficient editing ability is maintained in different cell types, and the target chain cutting mediated reverse pilot editing can be provided for the target chain cutting site. And meanwhile, the off-target risk is reduced. The srPE gene editing technology breaks through the fundamental limitation of the prior art, and provides a more comprehensive and more efficient technical tool for precise medicine and gene therapy.

The invention discloses application of an upstream non-coding region uORF of an ABA2 gene in regulation and control of crop agronomic traits, in particular to application in improvement of pre-harvest sprouting resistance of rice and cultivation of pre-harvest sprouting resistant rice strains, and belongs to the technical field of gene engineering. The invention finds that uORF in the upstream non-coding region of the crop ABA2 gene can inhibit the translation level of downstream ABA2 protein on the basis of not influencing the transcriptional level, so that the uORF can participate in regulation and control of crop agronomic traits in which ABA participates in regulation and control, such as improvement of pre-harvest sprouting resistance of rice. Therefore, the upstream non-coding region uORF of the crop ABA2 gene can be modified through a hybridization or gene editing technology, so that the purpose of regulating and controlling crop agronomic traits in which ABA participates in regulation and control is achieved, for example, site-directed mutagenesis is performed on an initiation codon of the upstream non-coding region uORF of the rice ABA2 gene, and the pre-harvest sprouting resistance of rice is improved.

The invention discloses a construction method and application of an NMD escape model of an Alport syndrome mouse. A non-human animal model carrying Col4a5 gene c.4432delG frame shift mutation is prepared on the basis of a CRISPR / Cas9 gene editing technology. The method comprises the following steps: co-injecting gRNA of a 49 exon of a targeted Col4a5 gene, homologous recombinant donor oligonucleotide containing c.4432delG mutation and Cas9 nuclease into a mouse fertilized egg, and carrying out embryo transplantation to obtain an F0-generation mutant mouse; a mutation site is verified by combining PCR (Polymerase Chain Reaction) with sequencing, and a stably inherited mutation line is established through two generations of breeding. Through verification, the model accords with pathological characteristics of the Alport syndrome, can stably simulate typical clinical manifestation and pathological characteristics of the human X-linked Alport syndrome, and can be used as an important tool for research of the Alport syndrome.

The invention belongs to the field of citrus breeding, and discloses application of a citrus CsERF113 gene in regulation and control of plant fruit abscission. It is found for the first time that the CsERF113 gene can induce plant separation layer formation and fruit abscission, which has a great promotion effect on molecular mechanism research of abscission regulation of plants including organs such as fruits and the like; the citrus CsERF113 gene cloned by the invention has a remarkable induction effect on citrus fruit abscission, which is of great significance to cultivation of overwintering non-abscission late-maturing citrus varieties through a gene editing technology.

The present invention provides a chimeric nucleic acid-guided nuclease including at least one domain substituted with distinct bacterial species. These chimeric nucleases improve function of modified guide polynucleotides, enhancing the genomic modification abilities of nucleic acid-guided nucleases. CRISPR-Cas systems such as those described herein are often used in genome editing, but the complexity of the polypeptides and polynucleotides leads to unpredictable outcomes. The chimeric CRISPR-Cas system disclosed herein was surprisingly able to rescue function of a previously non-functional guide nucleotide. The nucleases and methods described herein are useful for treating diseases associated with a gene or polynucleotide.

The invention provides a method for constructing a rice epigenetic recombinant inbred line, and belongs to the technical field of molecular biology and genetic breeding. According to the method for constructing the rice epigenetic recombination inbred line, a fertile rice DDM1 gene function weakening mutant is obtained through a CRISPR / Cas9 gene editing technology and hybridized with wild type rice with the same background to obtain F1, then the F1 and the wild type rice are subjected to backcrossing to obtain BC1F1, single-plant offspring selfing with the DDM1 genotype being the wild type is selected, selfing is conducted for multiple generations through a single-grain transmission method, and the rice epigenetic recombination inbred line is obtained. According to the present invention, the strain population with stable inheritance is constructed, such that the rice epigenetic recombinant inbred line is obtained, and has characteristics of rich phenotype, genetic and epigenetic variation so as to provide important population resources for batch identification and the like of important agronomic shape functional sites of rice.

The invention belongs to the technical field of gene editing, and discloses a base editor system and application. According to the invention, a heterologous peptide sequence is inserted into IscB protein, the obtained peptide embedded IscB is fused with deaminase or glycosylase to construct a base editor, and DNA base editing is carried out. According to the base editor system, the size limitation is overcome, and single AAV delivery is achieved; the editing efficiency is obviously improved; the broad-spectrum applicability is realized; the system optimization is more flexible; the method has efficient editing potential in vivo, and the off-target effect is reduced. The invention provides an important new strategy for hyperlipemia treatment and provides a basis for gene therapy of other hereditary diseases.

The invention relates to the technical field of gene editing, and particularly provides a method for creating herbicide-resistant rice through gene editing of an OsEPSPS gene promoter region, and the method is characterized in that WT / 29bp single allelic deletion mutation is introduced into the OsEPSPS gene promoter region, i.e., A in an OsEPSPS gene initiation codon ATG is 0 site, A in the OsEPSPS gene initiation codon ATG is 0 site, A in the OsEPSPS gene initiation codon ATG is 0 site, and A in the OsEPSPS gene initiation codon ATG is 0 site; 29bp is deleted from the 112nd site to the 1150th site at the upstream of the initiation codon ATG on one allele, and the other allele is kept unchanged. By introducing the single allelic deletion mutation, the 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, and the application value is high for guaranteeing agricultural production safety and improving the use efficiency of the herbicides.

The invention provides an application of WNT7B in construction of a myopia animal model. Homologous genes wnt7ba and wnt7bb of the WNT7B in the zebra fish are knocked out or knocked down by applying a CRISPR / Cas9 gene editing technology, an animal model of which the eye axis length is remarkably increased, the eyeball movement frequency is remarkably reduced and the light response is reduced is obtained, and the result shows that the WNT7B is related to the high myopia. The model provides a favorable tool for pathogenesis and drug screening of high myopia, and has a good application prospect.

The invention belongs to the technical field of plant genetic engineering, and particularly relates to application of a wheat transcription factor TaWRKY22 and a coding gene thereof in regulation and control of plant stripe rust resistance. The function of the wheat transcription factor TaWRKY22 is researched through molecular biology and genetics methods, it is found that knockout of the TaWRKY22 gene significantly improves the resistance of wheat to stripe rust, overexpression of the TaWRKY22 gene reduces the resistance of wheat to stripe rust, and then it is indicated that the wheat transcription factor TaWRKY22 plays a negative regulation role in wheat stripe rust resistance. Therefore, the TaWRKY22 gene is subjected to targeted modification through a gene editing technology, a new genetic material and a molecular breeding strategy can be provided for wheat stripe rust resistance breeding, and an important material and technical support are provided for breeding of wheat disease-resistant varieties.

The invention discloses application of a gene CsPOD7 in cucumber haploid breeding, and belongs to the technical field of molecular breeding. According to the invention, a parthenogenesis haploid induction gene CsPOD7 is cloned from cucumber, the CsPOD7 is knocked out through a CRISPR-Cas9 gene editing technology, and a cspod7 mutant is used as a male parent to be hybridized with other cucumber materials as a female parent, so that parthenogenesis haploid can be generated. The invention is the first application of the gene CsPOD7 in developing a parthenogenesis haploid induction line in dicotyledon, realizes the application of the haploid induction line based on the gene CsPOD7 in cucumber genetic breeding, accelerates the cucumber breeding process, and has huge potential application value and market prospect.

The invention discloses a VpCas9 protein, a single site mutant of the VpCas9 protein and application of the VpCas9 protein in gene editing. According to the invention, mining and modification of Cas9 are carried out through a deep learning model, new Cas9 protein VpCas9 protein is finally obtained through screening, and the amino acid sequence of the Cas9 protein VpCas9 protein is shown as SEQ ID No.5; compared with the SpCas9 protein, the VpCas9 protein has a better editing effect in the aspects of gene editing efficiency, PAM compatibility or specificity and the like; according to the invention, single site mutants with higher editing efficiency are further obtained through sequence screening and optimization design, and compared with SpCas9 protein and VpCas9 protein, the single site mutants have a better gene editing effect in genome editing; the VpCas9 protein and the single site mutant thereof provided by the invention have application prospects in genome editing or target nucleic acid editing of plants, animals or microorganisms.

The invention provides a tumor cell vaccine as well as a preparation method and application thereof. The preparation method comprises the following steps: acquiring and culturing tumor cells; enabling the cell membrane of the tumor cell to express a targeting antibody by utilizing a gene vector infection or gene editing technology, and enabling the targeting antibody to be used for enabling the tumor vaccine to be combined with the dendritic cell; performing overexpression of tumor specific protein on the cell membrane surface of the tumor cell by using a gene vector infection or gene editing technology; a nucleic acid substance of a target protein is wrapped by lipid nanoparticles to form a vaccine framework, and the nucleic acid substance can enable target cells to express directional chemotactic molecules; wherein the directed chemotactic molecule is used for transferring the dendritic cells to lymph nodes; the target vaccine is obtained by wrapping the vaccine framework with the cell membranes of the tumor cells, lymph node homing of the dendritic cells can be promoted, and the antigen presentation efficiency is increased and the immune effect is enhanced through tumor specific protein carried by the target vaccine and assisted by an immunologic adjuvant.

The invention belongs to the technical field of gene editing, and particularly relates to optimized omega RNA and gene editing application thereof. According to the optimized omega RNA, base mutation is carried out on the basis of wild type omega RNA with a gene sequence shown as SEQ ID NO.1, and by taking the 5'end of the wild type omega RNA, the 36th paired base UA is replaced with GC, the 37th paired base UA is replaced with AU, the 38th paired base UG is replaced with AU, the 50th paired base UG is replaced with CG, and the 51st paired base UA is replaced with GC. Compared with wild type omega RNA, the omega RNA has the advantages that the maximum enhancement multiple of the insertion and deletion activity is 16.7 times, 15 pairs of paired bases from the 40th site to the 54th site of the omega RNA are further deleted, and the editing efficiency is further improved.

The invention discloses a male sterility gene ZmUCE4 and application thereof in creating a male sterility line of corn, and belongs to the field of plant biotechnology breeding. According to the invention, site-directed mutagenesis of the ZmUCE4 gene in wild type corn is carried out through a CRISPR / Cas9 gene editing technology, pollen development abnormality and abortion can be caused, complete male sterility is caused, and it is proved that the ZmUCE4 gene has a regulation and control function on corn male reproductive development. Through offspring screening, a sterile line without transgenic components can be obtained, a stable maize male sterile line is created, and the method has important significance on maize male fertility control and hybrid seed production. The invention also designs a functional molecular marker aiming at the obtained uce4 male sterility mutant gene, and has important application value in maize male sterility line cultivation, sterile hybrid seed production and molecular marker-assisted selection.

The invention belongs to the technical field of genetic engineering, and particularly relates to a method for regulating and controlling production of human milk oligosaccharide through membrane protein. The method is realized by inactivating a membrane protein YgfX coding gene on a production strain. According to the human milk oligosaccharide production strain obtained in the mode that the membrane protein YgfX is inactivated after the coding gene of the membrane protein YgfX is knocked out or deleted from a 192nd guanine base through a gene editing technology, the human milk oligosaccharide production capacity of the human milk oligosaccharide production strain is obviously improved compared with that before gene editing, the yields of LNTII, LNnT, LNT, 3 '-SL and 6'-SL are improved by about 40%, 125%, 46%, 32% and 47% respectively, the effect is remarkable, and the human milk oligosaccharide production strain has a good application prospect. And a new method and thought are provided for widening the production way of the human milk oligosaccharide.

The invention discloses a VpCas9 protein double-site mutant and an application of the VpCas9 protein double-site mutant in gene editing. The invention firstly provides a homologue VpCas9 protein of Cas9, the amino acid sequence of the homologue VpCas9 protein is shown as SEQ ID No.5, and compared with SpCas9, the homologue VpCas9 protein has a better editing effect in the aspects of gene editing efficiency, PAM compatibility or specificity and the like; in order to further improve the gene editing effect of the VpCas9 protein in genome editing, the VpCas9 protein is subjected to double-site mutation, the VpCas9 protein double-site mutant is obtained, and compared with SpCas9 protein or VpCas9 protein, the VpCas9 protein double-site mutant has a better gene editing effect in genome editing, and the gene editing effect of the VpCas9 protein is improved. The method has an important application prospect in genome editing of plants, animals or microorganisms.

The invention discloses a method for establishing an animal model for age-related retinopathy caused by lipid metabolism disorder, and the animal model is obtained by specifically knocking out Lss genes from mouse retinal pigment epithelium (RPE) cells by means of a CRISPR / Cas9 gene editing technology. The age-related retinopathy model mouse constructed by the invention shows a retinopathy phenotype with retinal lipid deposition and reduced visual function at the age of 5 months, and the retinopathy is further aggravated along with the increase of the age of the mouse and is similar to the phenotype of age-related retinopathy related to lipid metabolism disorder clinically; and an ideal and effective animal model is provided for researching age-related retinopathy.

The invention belongs to the technical field of gene engineering, and provides rice purple acid phosphatase OsPAP16 and application thereof, and the application is that a rice purple acid phosphatase OsPAP16 mutant is applied to rice cultivation. Two OsPAP16 mutants are prepared by adopting a gene editing technology, then the mutants are subjected to overexpression induction, and the result shows that the purple acid phosphatase OsPAP16 is positioned on a cell membrane, the OsPAP16 is subjected to phosphorus deficiency induced expression on leaves and roots, the phosphorus deficiency induced expression at the roots is quicker and higher, the overexpression of the OsPAP16 can improve the activity of the acid phosphatase on the root surface, and the yield of the acid phosphatase on the root surface is increased. Phosphorus deficiency stress of the rice can be relieved by degrading organic phosphorus in the environment, the phosphorus content of the rice is increased, and then growth and development of the rice are promoted.

The invention relates to the field of plant genetic engineering, in particular to a rice OsDNG1 protein, a coding gene and application of the rice OsDNG1 protein to regulation and control of grain shape and grain weight. According to the invention, the OsDNG1 gene is knocked out through a CRISPR / Cas9 gene editing technology, so that the length of rice grains can be increased by 4.10%-4.62%, and the thousand grain weight can be increased by 10.86%-11.49%. And if the gene is over-expressed, the grain length is reduced by 1.14%-1.32%, and the thousand grain weight is reduced by 8.24%-8.56%. In the re-sequenced indica rice variety, the indica rice variety without the OsDNG1 gene type is longer in grain length and larger in length-width ratio compared with the variety with the gene type. The invention not only provides a key target for analyzing the molecular mechanism of rice grain morphogenesis, but also provides important theoretical basis and technical support for molecular design breeding of rice grain shape and grain weight, and has remarkable agricultural application value.

The invention provides a genome editor enDelIscB and a genome editor enDelIscB-T5E based on an IscB system, and further provides a base editor ICBE and a base editor IABE based on the enDelIscBD60A. According to the invention, a genome editor and a base editor which are more compact in size and higher in cleavage activity and insertion / deletion efficiency are realized by optimizing a single mutation site of the activity of a DelIscB protein cleavage plasmid, an optimized variant of the DelIscB protein and an optimized sgRNA variant of the DelIscB. According to the present invention, it is verified that the albinism model is efficiently constructed by using the enDelIscB editor and the enDelIscB-T5E editor through micro-injection of the mouse embryo, such that the enDelIscB editor and the enDelIscB-T5E editor have great potential in the field of genome editing.

The invention is applicable to the technical field of rabbit breeding, and provides a method for increasing the body weight of a rabbit based on a gene editing technology, which comprises the following steps: designing an action target sgRNA on a rabbit NR6A1 gene aiming at an NR6A1 p.P192S site; uniformly mixing the synthesized sgRNA and SpRY-CBE, injecting fertilized eggs, transplanting the fertilized eggs into fallopian tubes of receptor female rabbits, and feeding the receptor female rabbits until production; dNA is extracted from produced young rabbit ear tissues, PCR and sequencing are carried out, and if C at the fifth site in NR6A1 gene target sites is mutated into T, it is proved that single base mutation is obtained, and the NR6A1 gene edited rabbit is obtained. According to the invention, the rabbit variety with increased weight is successfully cultivated; a new thought of animal genetic breeding is developed by utilizing a gene editing technology, the method has application values in promoting improvement of genetic characters of rabbits and improving production performance and economic value of the rabbits, and powerful support is provided for high-quality development of the animal breeding industry in China.