258 results about "Zygote" patented technology

The invention relates to an artificial hybridization fry breeding method for epinephelus fuscoguttatus and pink pink bass, which belongs to the technical field of distant hybridization breeding and comprises the steps of parent selection of epinephelus fuscoguttatus and pink pink bass, parent breeding, parent intensive breeding, gamete collection, artificial insemination, hatching of fertilized eggs and fry breeding. By means of reproductive regulation and distant hybridization technologies, two species belonging to two genera under the grouper family with different geographical distribution and different breeding time are hybridized, namely cross-generic hybridization under the grouper family is achieved, intergeneric reproductive isolation is broken through, and the breeding efficiency is improved. Therefore, new hybridized germplasm with excellent characteristics of high fertilization rate, high hatching rate, high survival rate, high growth speed and the like is cultivated.

The invention provides an enhancer RNA (Ribonucleic Acid) molecule MZGAe1 and application thereof, and relates to the technical field of biology. The invention provides an enhancer RNA (Ribonucleic Acid) molecule MZGAe1. The nucleotide sequence of the enhancer RNA molecule MZGAe1 is as shown in SEQ ID NO: 1; meanwhile, the invention further provides a specific sgRNA sequence for activating the molecule in a targeted manner and a recombinant vector of the specific sgRNA sequence. Expression of endogenous MZGAe1 of cells is specifically activated by adopting a CRISPR activation technology, and conversion of mouse embryonic stem cells to bicellular-like cells can be efficiently promoted. By providing a brand-new endogenous targeted MZGAe1 accurate activation tool, the efficient transformation of wild mouse embryonic stem cells to bicellular cells can be realized only under the condition of endogenous activation of single enhancer RNA molecule MZGAe1, the proportion is at least increased by 5%, the expression of a totipotent marker gene ZSCAN4 is activated, and the expression of the totipotent marker gene ZSCAN4 is promoted. And an efficient and specific brand-new research tool and scheme are provided for researching zygote genome activation, embryonic development early events and cell reprogramming.

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 discloses an image detection and evaluation method for an unfrozen form of a fertilized egg embryo. The method comprises the following steps: acquiring a time sequence image sequence after the fertilized egg embryo is unfrozen; performing image standardization processing on the time sequence image sequence to generate a standardized embryo image set; extracting multi-dimensional embryo quality parameters from the standardized embryo image set; wherein the multi-dimensional embryo quality parameters comprise a morphological static parameter set, a dynamic change parameter set and a texture feature parameter set; generating a blastocyst formation rate based on the multi-dimensional embryo quality parameters; outputting a third-level embryo quality evaluation result according to a comparison relationship between the blastocyst formation rate and a preset threshold interval; wherein the three-stage embryo quality evaluation result comprises an excellent embryo grade, a qualified embryo grade and an abnormal embryo grade. The method has the following advantages and effects: full-automatic and multi-dimensional quantitative evaluation of the thawing process of the unfrozen embryos is realized, so that the scientificity of embryo screening and the success rate of clinical pregnancy are improved.

The invention discloses an artificial cultivation method of a hybrid of epinephelus trifoliatus and epinephelus lanceolatus, which comprises the following steps: taking healthy epinephelus trifoliatus as female parent fish, and carrying out cultivation strengthening and artificial ripening to obtain strengthened female parent fish; healthy epinephelus lanceolatus is used as male parent fish, and sexual mature male parent fish is obtained through natural sexual maturity; the ovulation time and the spermiation time are consistent; collecting egg liquid and seminal fluid, inseminating to obtain fertilized eggs, and performing embryo incubation and illumination regulation and control cultivation after opening to obtain hybrid spots; carrying out authenticity identification on the hybrid spots by utilizing a primer pair, and determining that the hybrid spots inherit genetic information of the female parent fish and the male parent fish; the primer pair is a sequence pair composed of SEQ ID NO. 1 and SEQ ID NO. 2, and a sequence pair composed of SEQ ID NO. 3 and SEQ ID NO. 4. The technical blank of crossbreeding by taking the epinephelus trifoliatus as the parent epinephelus is filled, and the authenticity of the hybridized spots is reliable.

The invention relates to the field of distant hybridization breeding, and discloses an artificial breeding method for hybrid species of humpback perches and epinephelus fuscoguttatus, which comprises the following steps: selecting healthy humpback perches as female parent fishes, and screening healthy epinephelus fuscoguttatus as male parent fishes; carrying out intensive cultivation on the parents by means of nutrient enrichment and temperature regulation and control, and promoting synchronous development and maturation of gonads of the parents; after the parents reach the breeding state, eggs of the female parent fishes and seminal fluid of the male parent fishes are collected respectively, fertilized eggs are obtained through artificial insemination, incubation management such as temperature control and continuous oxygenation is conducted on the fertilized eggs, and finally hybrid grouper fries are obtained. The crossbreeding application of the humpback perch as the female parent and the epinephelus fuscoguttatus is realized, and the problems that in the traditional hybridization process, parent gonad development is asynchronous, and filial generations cannot develop are effectively solved; the bred hybrid provides technical guarantee for standardization and precision promotion of grouper crossbreeding, and creates favorable conditions for breeding new hybrid lines with excellent characters of parents.

The invention provides a breeding method of hypoxia-resistant golden tiger hybrid spots, which comprises the following steps: carrying out artificial insemination on a female parent of epinephelus coioides and a male parent of epinephelus coioides to obtain fertilized eggs, carrying out 2.8-3.2 mg / L hypoxia screening on the female parent for more than three generations, and ensuring that the suffocation point is less than or equal to 0.9 mg / L. Performing staged low-oxygen incubation on the fertilized eggs, gradually reducing dissolved oxygen in an embryonic period, a protointestinal period and a membrane emergence period, performing specific sound wave stimulation in an opening period of larva fish, performing low-oxygen-resistant phenotype screening in a larva period, performing intermittent low-oxygen intensive training in a juvenile fish period, applying water flow stimulation before training, and performing low-oxygen-resistant phenotype screening in a membrane emergence period; during the period, a fortified feed containing gamma-aminobutyric acid, heme iron and other components is fed. According to the method, through multi-link collaborative innovation, the hypoxia resistance of the Jinhu hybrid spots is effectively improved, the survival rate of a target group under 2.5 mg / L dissolved oxygen reaches 93%, the suffocation point is reduced to 0.75 mg / L, meanwhile, gill filament development and growth are promoted, and an efficient technical scheme is provided for aquatic product breeding.

The invention relates to the technical field of fishery breeding, and particularly discloses a fry hatching device for schizothorax breeding, comprising a support frame, the top of the support frame is fixedly connected with a hatching box and a breeding box, the side surface of the hatching box is fixedly connected with the side surface of the breeding box, and two sides of the inner wall of the breeding box are fixedly connected with plastic plates; the fry hatching equipment comprises a hatching box and a breeding box, the side faces of the hatching box and the breeding box communicate with water adding pipes, the bottom of the side face of the hatching box penetrates through and is fixedly connected with a purification assembly, and an inlet is formed in the side, close to the hatching box, of the inner wall of the breeding box. And the aeration device is arranged, so that the content of dissolved oxygen in the incubator can be increased, sufficient oxygen is provided for fertilized eggs, and healthy growth of the fertilized eggs is promoted.

The invention provides a method for overspeed preparation of gene edited goldfish. The method comprises the following steps: S1, obtaining gobiocypris rarus without primordial germ cells as receptor fish; s2, designing a target spot for the goldfish amh gene, introducing a Cas9 / gRNA complex into a goldfish fertilized egg, performing sequencing verification to obtain a mutant individual, and feeding the mutant individual as a donor fish; s3, taking gonads of donor fish, cutting into pieces, performing enzymolysis digestion, filtering, and performing density gradient centrifugation to enrich donor germline stem cells; s4, transplanting the germline stem cells of the donor fish into the receptor fish body, and feeding to obtain the gene edited goldfish sperms and ova. S5, hybridizing and feeding goldfish amh mutant sperms and ova produced by gobiocypris rarus to obtain gene edited goldfish individuals. According to the invention, gobiocypris gobiocypris is used as a receptor, the cross-subfamily gene edited goldfish is used as a donor, and an efficient, stable and easy-to-popularize method for preparing the gene edited goldfish is provided by utilizing a belly borrowing reproduction technology.

The invention provides an artificial propagation method of Sinilabeo wuyi, and belongs to the technical field of aquatic animal breeding. The artificial propagation method of the Sinilabeo wuyi comprises the steps of parent fish classification, parent fish rearing, artificial spawning induction, natural insemination, artificial insemination, fertilized egg incubation, fry rearing, juvenile fish rearing and the like. The oxytocic is reasonable in compatibility, the injection time of the oxytocic is accurate, the oxytocic rate, the fertilization rate and the hatching rate are increased, and damage to parent fishes is small. According to the method, feeding methods in different stages of fry breeding are also defined, the survival rate of the sinilabeo wuyi fry breeding stage is increased, and a technical support is laid for artificial breeding of the endangered species sinilabeo wuyi.

The invention belongs to the field of fish germplasm improvement and innovation, and discloses an application of river perch sperms in inducing gynogenesis of largemouth bass, which comprises the following steps: obtaining mature largemouth bass ova and river perch sperms through artificial spawning induction; the method comprises the following steps: collecting river perch semen, diluting, oscillating, irradiating and inactivating with an ultraviolet lamp, mixing with mature largemouth bass ova, inseminating, and hatching fertilized ova to obtain gynogenesis largemouth bass. According to the method, the sperms of the river perches are used for stimulating gynogenesis of the micropterus salmoides for the first time, the gynogenesis micropterus salmoides is successfully obtained, and the heterologous sperms in the far geographic space are used as the stimulation source of the artificial gynogenesis ovum for the first time. The method not only provides an innovative germplasm resource basis in the aspect of improving germplasm of the micropterus salmoides, but also has important significance in the research aspects of fish genetic breeding and'micro-hybridization among fish species in far geographic space '.

The invention belongs to the field of fish breeding, and discloses a construction method and primers of a fast-growing red crucian carp homozygous strain, and the construction method comprises the following steps: preparing gRNA of a red crucian carp acvr2b gene, microinjecting a mixture of the gRNA and Cas9 protein into a one-cell stage fertilized egg of the red crucian carp, and screening to obtain an F0-generation gene knockout mutant; carrying out selfing on the F0-generation gene knockout mutant to obtain an F1-generation mutant, and screening to obtain a homozygous mutant; and selfing and propagating the homozygous mutant to obtain the product. The acvr2b homozygous knockout strain is successfully established in the red crucian carp through a CRISPR / Cas9 gene editing technology for the first time, the body weight of the obtained acvr2b mutant red crucian carp at the age of 12 months is increased by 30.68% compared with that of a wild type, the muscle protein content is increased by 13.81%, the intestinal digestion and absorption capacity is obviously improved, high-quality germplasm resources are provided for fish genetic breeding, and the acvr2b mutant red crucian carp has a good application prospect.

The invention provides a full-indoor artificial breeding method for plectropomus leopardus, and belongs to the technical field of artificial breeding of marine fishes. The full-indoor artificial breeding method for the plectropomus leopardus comprises the steps of earlier-stage incubation of fertilized eggs, incubation in an indoor breeding pond, breeding in an indoor flowing water pond, breeding in an indoor circulating water pond and the like. By regulating and controlling illumination, the irritability of the fries is greatly reduced, the ecological conditions of the fries are effectively optimized, and the health condition and adaptability of the fries are improved. Meanwhile, different types and different total amounts of biological bait combinations are fed in the larva stage, the juvenile stage and the fish stage of the plectropomus leopardus, transition is made in each bait transfer period, enteritis caused by excessive intake of larvae and juveniles and too little intake caused by inadaptation of bait transfer are effectively reduced, and the problem that the survival rate is low in the bait transfer period can be effectively solved.

The invention belongs to the field of fish genetic breeding, and discloses a method for efficiently obtaining homozygous red crucian carp, which comprises the following steps: (1) carrying out artificial false fertilization on eggs of the red crucian carp to obtain fertilized eggs; (2) performing gene editing on the fertilized eggs; (3) incubating the fertilized eggs subjected to gene editing for 18-32 minutes, and then carrying out cold treatment; and (4) incubating the fertilized eggs after cold treatment to obtain the homozygous red crucian carp. According to the method disclosed by the invention, gene editing and gynogenesis methods are comprehensively utilized, the hatching rate, the survival rate and the homozygous rate are high while the high gene editing efficiency is maintained through the early cold shock treatment time and the proper cold shock treatment time, and the gynogenesis red crucian carp mutant with a high proportion can be obtained.

The present invention addresses the problem of specifically clarifying the effects on a fertilized egg when 5-ALA is administered to a human female suffering from infertility, and constructing a more effective means for treating infertility through the suppression of chromosomal abnormalities. It has been confirmed that, when 5-ALA is ingested in combination with a drug therapy that is generally performed in the treatment of female infertility, the rate of chromosome aneuploidy in a fertilized egg or the like is significantly reduced, and the qualitative improvement of an embryo through improvement of the stability of the chromosomes is expected.

The application discloses a submerged Cinnamomum micranthum in-vitro rapid propagation and brown-preventing seedling raising method based on zygote embryos, directly induces adventitious buds of the submerged Cinnamomum micranthum for the first time, significantly improves a propagation coefficient, and overcomes problems such as vitrification and browning of the adventitious buds of the submerged Cinnamomum micranthum. The application establishes an efficient and stable in-vitro regeneration system of the submerged Cinnamomum micranthum, and provides a basis for large-scale seedling raising of the submerged Cinnamomum micranthum.

The invention provides a culter turunx2b gene knockout method and application of the culter turunx2b gene knockout method in creation of intermuscular thorn-free mutants, and belongs to the technical field of gene editing. The invention provides an efficient gene knockout method which comprises the following steps: selecting two targets on a second exon of a culter turunx2b gene as target sites, designing and synthesizing two gRNAs, mixing the gRNAs with Cas9mRNA, introducing the mixture into a fertilized egg of culter turunx2b in an I cell stage in a microinjection manner, hatching, and screening mutated F0-generation culter turunx2b individuals. By utilizing the method disclosed by the invention, the F0-generation knockout efficiency is up to 87.5% under the condition of ensuring that the fertilized eggs are hatched to survive. The method disclosed by the invention is simple and easy to implement and simple to operate, the culter alburnus gene mutation strain can be rapidly obtained by utilizing the gene mutation method, and the method has important scientific significance for developing economic fish gene function research, revealing genetic development law and creating new germplasm.

The invention discloses a bactrocera dorsalis point mutation gene editing method based on CRISPR-Cas9 and homologous directional repair, and relates to the technical field of gene editing. The method comprises the following steps: designing high-specificity sgRNA for a PAM sequence adjacent region of a target gene; extending the length of the homologous arm to 100bp, and designing a single-stranded DNA (deoxyribonucleic acid) containing a mutation site as a homologous repair template; after a mixture containing the high-specificity sgRNA, the homologous repair template and Cas9 protein is injected into a bactrocera dorsalis fertilized egg in a microinjection mode, incubation and incubation are carried out, and G0-generation imagoes are obtained through culture; hybridizing the G0-generation imago with wild bactrocera dorsalis, and screening to obtain a G1-generation hybrid mutant; and carrying out selfing by using the G1-generation heterozygous mutant, and screening to obtain a G2-generation homozygous mutant. By adopting the method, the mutation efficiency and embryo survival rate of bactrocera dorsalis can be effectively improved.

This invention discloses a gRNA combination and its application in the preparation of drugs for the prevention of metabolic-associated fatty liver disease (MASLD). The gRNA combination, when mixed with Cas9 protein, yields an RNP complex, which, when microinjected into mouse zygotes, can breed a stable and heritable mouse strain with TMEM68 gene knockout. Combining lipidomics, transcriptomics, and primary hepatocyte functional verification, the core regulatory role of TMEM68 in hepatic lipid metabolism is revealed for the first time systematically. Experiments demonstrate that TMEM68 deficiency significantly reduces the storage of triglycerides (TAG) in the liver and hepatocytes, decreases lipid droplet formation, and reshapes the metabolic homeostasis of various lipids such as glycerophospholipids, cholesterol esters, and bile acids. Therefore, the gRNA combination of this invention, or the RNP complex obtained by mixing the gRNA combination with Cas9 protein, can be used to prepare drugs for the prevention of MASLD, providing a novel intervention strategy for MASLD prevention with broad application prospects.

The invention belongs to the technical field of GPCR drug research and development, and discloses a ratio type cAMP fluorescent probe RaCamp and a construction method of a transgenic mouse, the probe RaCamp is formed by series fusion of G-Flamp2 fluorescent protein and mCherry fluorescent protein through a flexible linker, and the preparation method of the probe RaCamp comprises the following steps: S1, obtaining an mCherry fragment; s2, a G-Flamp2 linear carrier is obtained; s3, homologous recombination; s4, converting and screening; the construction method of the transgenic mouse comprises the following steps: T1, designing a carrier; t2, microinjection of fertilized eggs; t3, embryo transplantation and reproduction; the invention has the beneficial effects that mCherry is added as an internal reference protein on the premise of keeping the sensitivity of the original G-Flamp2 probe, and the selected connecting peptide (GGGGS) 2 enables the expression quantity of the two proteins in cells to be accurately controlled to be 1: 1, so that the problem of background interference in cell observation of in-vivo imaging is well solved. And the sequence is inserted into a Rosa26-LSL box, so that the construction of a Rosa26-LSL-RaCamp mouse is realized.

The name of the invention is NK zygote molecules and methods of use thereof. Compositions for activating NK cells to stimulate an immune response for the treatment of cancer and other disorders are provided. In one embodiment, the present invention provides a compound comprising an NK binding domain that binds to CD16; an NK activation domain operably linked to the NK binding domain; and a targeting domain selectively binding to a target cell and operably linked to the NK activating domain and the NK binding domain, wherein the targeting domain binds to CLEC12A.

The application relates to the field of aquaculture, and particularly discloses a method for cultivating oyster larvae, which comprises the following steps: S1, fertilized eggs are developed into D-shaped larvae in seawater at 22 DEG C for 20-24 hours; S2, the D-shaped larvae are cultivated under the condition that the temperature is 22-24 DEG C and the salinity is 29-31 ‰, feed and growth nutrient solution are fed daily, water is changed daily, after the D-shaped larvae appear eye spots, a larval collector is put in, and algal feed and growth promoting solution are fed daily, the larvae are cultivated into 0.8-1 mm juvenile shellfish, and are transferred to sea cultivation; the growth nutrient solution comprises the following components in weight parts: 1-2 parts of a small molecule peptide solution, 0.01-0.03 parts of neomycin sulfate powder, 0.5-1 part of corn starch, 2-4 parts of modified montmorillonite, and 10-15 parts of water. The method for cultivating oyster larvae has the advantages that the survival rate of the larvae is improved, the growth speed is accelerated, and the metamorphosis and attachment rate is improved.

The invention relates to the technical field of gene engineering, and discloses a construction method of a humanized zebrafish amyotrophic lateral sclerosis model. Comprising the following steps: constructing a knock-in donor plasmid containing a left homologous arm sequence, a human mutation SOD1 gene sequence and a 3 'UTR sequence of a zebra fish sod1 gene, wherein the left homologous arm contains a target sequence of sgRNA and a second exon sequence of the zebra fish sod1 gene; the method comprises the following steps: knocking in a donor plasmid and Cas9 protein into sgRNA of which the target sequence is as shown in SEQ ID NO: 1, injecting the donor plasmid and the Cas9 protein into fertilized eggs of zebra fish, culturing into adult fish, and screening out a strain of which the human mutation SOD1 gene is correctly inserted into a zebra fish genome. According to the invention, sgRNA with a specific sequence is matched with a corresponding method, so that the human mutated SOD1 gene can be inserted into a zebra fish genome, the expression of the human mutated SOD1 is realized, the expression of an endogenous sod1 gene in the zebra fish is destroyed, and the construction of a humanized ALS model is realized for the first time.

The invention provides a hematopoietic anti-aging active component extracted from fish embryos as well as an extraction method and application of the hematopoietic anti-aging active component, and belongs to the technical field of active component extraction. The hematopoietic anti-aging active component is extracted from fish embryos which are developed from fish fertilized eggs to the earlier stage of incubation at the optimum temperature and stop development through rapid freezing, and is rich in exosomes, active protein, peptide, chondroitin sulfate, hyaluronic acid, small molecule metabolites, various active factors and the like, and all the nutritional active components are matched with one another, so that the effects of resisting aging and resisting aging are achieved. The cartilage development can be supported, the joint health is ensured, the hematopoietic effect is enhanced, and the skin aging is delayed. The orally disintegrating tablet containing the hematopoietic anti-aging active component can avoid inactivation of bioactive substances caused by high temperature, and the biological activity of the hematopoietic anti-aging active component is reserved to the maximum extent. The hematopoietic anti-aging active component and the orally disintegrating tablet are simple in preparation process and remarkable in hematopoietic anti-aging effect.

The application belongs to the technical field of fish genetic breeding, and specifically discloses an artificial induction method of Megalobranchius ambius and Culter alburnus hybrid triploid and application. The artificial induction method comprises the following steps: S1. taking Megalobranchius ambius as a female parent and Culter alburnus as a male parent to perform artificial dry insemination; S2. performing hydrostatic pressure treatment on the fertilized eggs, and the hydrostatic pressure treatment conditions are as follows: the starting time of treatment after fertilization is 2-3 min, the pressure is 35-50 MPa, and the duration is 3 min; S3. hatching the fertilized eggs and performing transfer culture, and the water temperature in the whole process from fertilization, hydrostatic pressure treatment, hatching to fry cultivation is controlled to be 22-24 DEG C; and Megalobranchius ambius and Culter alburnus triploid is screened through ploidy analysis. The method can successfully induce Megalobranchius ambius and Culter alburnus hybrid sterile triploid with superior growth performance, and based on comprehensive index evaluation, the best hydrostatic pressure treatment condition is obtained, high triploid rate (>80%) and high survival rate (>98%) are simultaneously achieved, and commercialized breeding production can be realized.

The application provides a method for on-site rapid artificial insemination of wild Pseudosciaena crocea of Lutjanidae, and belongs to the technical field of aquatic animal breeding. Wild parent fish of Pseudosciaena crocea of Lutjanidae are captured in the breeding season; sperm is collected from the selected sexually mature male parent fish on the capture site; after artificial sperm collection, sexually mature female parent fish are selected, and fish eggs are collected; high-quality fertilized eggs are obtained through artificial insemination; the artificial insemination comprises the following steps: adding sperm into fish eggs, adding natural seawater for stirring, and finally washing with natural seawater. Through comprehensive regulation of key links in the whole process of artificial insemination, the purpose of obtaining high-quality fertilized eggs by using wild parent fish of Pseudosciaena crocea of Lutjanidae and carrying out artificial fry breeding is achieved.

The invention discloses an artificial breeding method of lota lota, which comprises the following steps: S1, selecting robust lota lota as parent fish, placing the parent fish in an indoor culture tank for temporary culture, and inducing gonad to develop mature by regulating and controlling water temperature; s2, performing artificial spawning induction on the parent fish with mature gonad to obtain ova and semen, and performing in-vitro fertilization on the ova and the semen to obtain fertilized ova; s3, performing two-step soaking treatment on the fertilized eggs by using a composite biological functional liquid containing a liquid A and a liquid B, and then transferring the fertilized eggs into a hatching facility for hatching to obtain fish fries; and S4, breeding the hatched fish fries. According to the artificial breeding method of the lota lota, the survival rate of fertilized eggs and the survival rate in the fry breeding stage are increased to a high level, large-scale and controllable artificial breeding of the lota lota becomes possible, and the overall technical scheme has high stability and industrial application value.

Methods for improving parthenogenesis efficiency by DWT1 and BABY BOOM transcription factors in plants are provided. A rice embryo trigger transcription factor BABY BOOM1 can initiate embryogenesis when expressed in the unfertilized egg cell through a process called parthenogenesis (Khanday et al., 2019. Nature 565: 91-95). The parthenogenesis efficiency by BABY BBOM1 itself is 10-29%. This invention describes methods of high frequency of parthenogenesis by simultaneous expression of BABY BOOM and DWT1 transcription factors. When BABY BOOM1 and DWT1 are expressed together through egg cell-specific promoters, parthenogenesis efficiencies of up to 90% are achieved. These high parthenogenesis efficiencies are a prerequisite for field applications of synthetic apomixis in crop plants.

The invention discloses a construction method of a COL4A5-K229X point mutation X-linked Alport syndrome mouse model. The construction method comprises the following steps: aiming at c.685Agt of a No.12 exon of a mouse COL4A5 gene; carrying out T point mutation, and designing and preparing Cas9 mRNA, gRNA and a donor vector; the components are mixed and then microinjected into fertilized eggs of a C57BL / 6J mouse to obtain an F0-generation mouse; identifying the genotype through PCR (Polymerase Chain Reaction) amplification and Sanger sequencing, and screening positive mice; mating the positive F0-generation mice with the wild-type mice, and breeding F1-generation and subsequent generations; the phenotype of the model is further verified through qPCR, biochemical analysis, light microscopic examination, transmission electron microscope and immunofluorescence. The model constructed by the invention shows hematuria, proteinuria, azemia, podocyte loss and irregular thickening and layering of glomerular basement membrane, is consistent with phenotypes of human XLAS patients, and provides an animal model tool for analyzing pathogenesis and developing treatment strategies.

The invention discloses a construction method of a Klf6 gene knockout mouse embryo model, which comprises the following steps: S1, preparing a gene editing solution containing Cas9 enzyme and sgRNA for targeting a third exon region and a fourth exon region of a mouse Klf6 gene by utilizing a CRISPR / Cas9 gene editing technology; s2, taking out the fertilized eggs of the mouse, transfecting the fertilized eggs of the mouse with a gene editing solution, and then performing in-vitro culture on the fertilized eggs; s3, carrying out embryo transplantation on the two-cell stage embryo obtained in the step S2 into a pregnant female mouse, and then carrying out conventional feeding; and S4, 7-8 days after embryo transplantation, taking out the uterus and stripping the embryo to obtain the Klf6 gene knockout mouse embryo model. According to the method, gene knockout is carried out by using CRISPR / Cas9 in the fertilized egg period in combination with an electrotransfection method, and compared with an existing method, the construction process time can be greatly shortened, the working efficiency is obviously improved, and the success rate is high.