338 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 ecology-imitating artificial propagation method for Dongquasichthys nobilis. The ecology-imitating artificial propagation method comprises the following steps: S1, selecting male and female parent fishes and putting the male and female parent fishes into an artificial ecology-imitating breeding pond for breeding; s2, in a non-breeding season, feeding female parent fish feed A for 2-3 times every day, and feeding male parent fish feed B for 2-3 times every day; s3, 2-3 months before breeding, female parent fish fortified feed I is fed 2-3 times every day, and male parent fish fortified feed II is fed 2-3 times every day; s4, after the gonad of the parent fish is mature, artificial spawning induction is conducted, and fertilized eggs are collected and hatched. According to the method, the ecology-imitating breeding environment is manufactured, so that the east astrostroma parent fish adapts to the artificial environment, the irritability is reduced, the east astrostroma parent fish is fed with different feeds in the non-breeding season and the breeding season, the ecological factor and nutritional requirements for gonad development of the east astrostroma can be met, the immunity of the east astrostroma parent fish is improved, gonad maturation is promoted, and the survival rate of the east astrostroma parent fish is increased. The quality of sperms and ova is improved, the fertilization probability is improved, the malformation probability of hatched larvae is reduced, and the breeding efficiency is improved.

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 method for targeted creation of a fast-growing new tilapia strain, and belongs to the technical field of fish cultivation. The method specifically comprises the following steps: (1) taking a fragment, as shown in SEQ ID NO.1, on a third exon of a pomc gene as a gRNA target sequence; (2) designing and synthesizing a gRNA primer sequence; (3) amplifying by adopting the gRNA primer sequence and the gRNA plasmid, and then transcribing and purifying to obtain a gRNA fragment; (4) transferring the gRNA fragments into fertilized eggs of tilapia mossambica through gene editing to obtain F0-generation positive fish; and (5) hybridizing the F0-generation positive male fish with the wild-type female fish, screening out F1-generation parent fish, and culturing the F1-generation parent fish to obtain F2-generation tilapia with homozygous deletion mutation. According to the invention, the pomc homozygous knockout line is established in the Nile tilapia mossambica for the first time, the new fast-growing line is a complete gene function deletion model, the cost is lower, the efficiency is higher, and the phenotype can be stably inherited.

The invention provides auxiliary reproduction test tube infant fertilized egg culture equipment which comprises a constant-temperature bottom box and a sealing top box covering the top of the constant-temperature bottom box in a sealing mode, a culture dish cavity is formed in the middle of the top of the constant-temperature bottom box, and a variable culture dish with a variable culture plane is arranged in the culture dish cavity; compared with the prior art, when the fertilized eggs of the test tube baby are cultured, a proper amount of nutrient solution is put into the variable culture dish in a sterile laboratory, the fertilized eggs of the test tube baby are put into the nutrient solution to be cultured, and at the moment, the middle of the culture plane of the variable culture dish is concave downwards to be in a funnel shape; the test tube infant fertilized eggs are always located in the middle of the culture plane, cells start to split along with culture, the culture plane of the variable culture dish is gradually adjusted from a funnel shape to a plane shape along with the increase of the cells, and therefore the situation that the test tube infant fertilized eggs are excessively dispersed after being split is avoided, and stacking of multiple cells can be avoided; the observation condition is influenced.

The invention relates to a method for producing notoginsenoside and ginsenoside by utilizing panax notoginseng cotyledon embryos, and belongs to the technical field of plant adventitious root tissue culture. The method comprises the following steps: inducing embryogenic calluses by utilizing immature zygotic embryos of panax notoginseng; proliferating the panax notoginseng embryogenic callus on a plate culture medium; s2, putting the pseudo-ginseng embryogenic callus obtained in the step S2 into a liquid culture medium, carrying out cell dispersion, and sieving by using a sieve with the aperture of 24-30 meshes; a mechanical stirring type bioreactor is filled with a liquid culture medium, and the globular embryo and the heart-shaped embryo are placed in the mechanical stirring type bioreactor to be cultured; s3, recovering the filtrate in the step S3, filtering by using a screen with the aperture of 30-80 meshes and 80-150 meshes to obtain uniform panax notoginseng embryogenic cells, and differentiating the panax notoginseng embryogenic cells in a triangular flask for the step S4; and drying the cotyledon embryo obtained in the step S4 at 45 DEG C, and grinding into powder. According to the method, large-scale generation of cotyledon embryos can be completed only in 4-6 weeks, the production process is remarkably accelerated, the method is suitable for industrial batch production requirements, and the production efficiency is high.

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 a double-fluorescence-labeled engineering bacterium for in-situ tracing of horizontal transfer of plasmids in pseudomonas among bacteria as well as a construction method and application of the double-fluorescence-labeled engineering bacterium, pseudomonas is taken as an original strain, a green fluorescence strong expression component is integrated into a pseudomonas chromosome genome, and green fluorescence-labeled pseudomonas is obtained; a red fluorescence strong expression element is inserted into a multiple cloning site of a broad-host zygote, and is converted into the green fluorescence labeled pseudomonas to form a double-fluorescence labeled engineering bacterium. The double-fluorescence-labeled strain disclosed by the invention can generate corresponding green fluorescence or red fluorescence after being excited by a green or red fluorescence excitation wavelength; when the plasmids enter other bacteria under the action of conjugation or transformation, the recipient bacteria can only present red fluorescence under the excitation of the red fluorescence excitation wavelength, and the horizontal transfer condition of the plasmids in the pseudomonas bacteria among the bacteria under the in-situ condition can be accurately tracked through the intuitive method.

The invention relates to the technical field of grass carp (Ctenopharyngodon idella), in particular to a method for knocking out a grass carp SMYD3 gene and application of the grass carp SMYD3 gene. The nucleotide sequence of the gRNA is as shown in SEQ ID NO: 1. The method comprises the following steps: synthesizing gRNA of a target gene through in-vitro transcription, then mixing the gRNA with Cas9 protein, introducing the mixture into an animal pole in a grass carp fertilized egg single-cell period by adopting a microinjection mode, and hatching to obtain the grass carp F0 generation with the target gene mutation. According to the application, the knockout of the grass carp gene is efficiently realized, and an effective technical means is provided for developing grass carp breeding and gene function research.

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 a method for inducing ostrea rivularis triploid, and relates to the technical field of shellfish genetic breeding. The specific preparation method comprises the following steps: firstly, obtaining egg liquid and seminal fluid from gonads of female and male ostrea rivularis individuals, cleaning and curing the egg liquid, and adding the seminal fluid into the egg liquid after cleaning to obtain a fertilized egg solution; when the proportion of the first polar bodies in the eggs of the fertilized egg solution is 30-50%, adding an inducer into the fertilized egg solution for induction and cleaning to obtain induced fertilized eggs; and finally, hatching and screening the induced fertilized eggs to obtain D-shaped larvae, and culturing to obtain the crassostrea rivularis triploid population. The method for inducing the crassostrea rivularis triploid is high in triploid induction rate, high in final hatching rate, capable of stably surviving and good in application prospect.

The invention belongs to the technical field of gene engineering, and relates to a construction method of a mouse model capable of rapidly degrading MSTN protein, which comprises the following steps: sequentially inoculating a 5'homologous arm, 3xEAAAK, an FKBP1A gene synonymous mutation sequence, 3xEAAAK, a red fluorescent protein mCherry sequence and a 3 'homologous arm on a Donor vector to obtain a Donor recombinant vector; designing gRNA based on a third exon of the Mstn gene, mixing the gRNA, Cas9 protein and a Donor recombinant vector, injecting the mixture into a mouse fertilized egg, and performing in-vitro culture and transplantation to obtain an F0-generation mouse; and hybridizing the F0-generation mouse with a wild type, or continuously breeding the filial generation to obtain the knock-in mouse model. According to the invention, a dTAG targeted protein degradation technology is utilized, the obtained homozygous knock-in mouse is subjected to dTAG administration, and rapid and reversible degradation and removal of MSTN protein can be realized.

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 present invention relates to the field of nerve cell labeling methods and related animal model technologies, and specifically to a promoter of a lgals3bpb gene and its application in constructing a zebrafish model. The application of the promoter of the lgals3bpb gene in constructing a zebrafish model comprises the following steps performed in sequence: obtaining the promoter of the lgals3bpb gene through a PCR reaction; constructing an expression vector regulated by the promoter of the lgals3bpb gene; transferring the expression vector into zebrafish fertilized eggs, and obtaining a stable genetically modified zebrafish strain through culture, screening and multi-generation hybridization. This technical solution can solve the technical problem of the prior art due to the lack of a zebrafish animal model capable of locating and operating microglia, and creates conditions for studying microglia, conducting disease mechanism research, drug screening and drug efficacy testing, and has good application prospects.

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 discloses a construction method and application of a GSDMB intestinal epithelium specific knock-in mouse model. The construction method comprises the following steps: (1) designing gRNA according to an ROSA26 gene, constructing a Donor vector containing a GSDMB gene, introducing the gRNA and the Donor vector and Cas mRNA into a fertilized egg of a receptor mouse, transplanting the fertilized egg and Cas mRNA into a pseudopregnant mouse body for reproduction, and screening out a GSDMB gene knock-in F0-generation mouse; (2) mating and breeding the F0 generation mouse and a wild type mouse, and screening out an F1 heterozygote mouse with GSDMB knocked in; (3) carrying out inbreeding propagation on the F1 heterozygote mouse, and screening out an F2 homozygote mouse in which GSDMB is knocked; (4) the F2 homozygote mouse and the intestinal epithelium specific Vilin-Cre transgenic mouse are subjected to mating breeding, and a heterozygote mouse with GSDMB intestinal epithelium specific knock-in is screened out; and (5) carrying out inbreeding propagation on the heterozygote mouse with the GSDMB intestinal epithelium specific knock-in, and screening out the homozygote mouse with the GSDMB intestinal epithelium specific knock-in.

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 provides a method for on-site rapid artificial insemination of wild sal-family large yellow croakers, and belongs to the technical field of aquatic animal breeding. According to the method, wild parent fishes are captured in the breeding season of the sal-family pseudosciaena crocea; selecting sexually mature male parent fishes on a capturing site, and collecting seminal fluid for later use; after manual semen collection is finished, female parent fishes with sexual maturity are selected, and fish eggs are collected; performing artificial insemination to obtain high-quality fertilized eggs; the artificial insemination comprises the following steps: adding seminal fluid into the roes, then adding natural seawater, stirring, and finally washing with the natural seawater. According to the method, key links of the whole artificial insemination process are comprehensively regulated and controlled, so that the purposes of obtaining high-quality fertilized eggs and carrying out artificial fry breeding by utilizing the parent fish of the wild sal ammoniac large yellow croaker are achieved.

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 invention provides a breeding method of fast-growing high-quality Dongzhihuo. Comprising the following steps: identifying the genotype of an SNP (Single Nucleotide Polymorphism) locus which is located at the 79th site of a gene sequence as shown in SEQ ID NO.1, and if the genotype of the locus is TT type, determining that the locus is a candidate parent; selecting candidate parents with uniform and robust body types as parent fishes; carrying out spawning induction on the male fish and the female fish of the Dongstellate star; collecting ova and sperms for artificial insemination; uniformly putting the fertilized eggs into an indoor nursery pond for hatching and breeding. The euryale ferox bred through the method is high in growth and development speed, high in muscle fat content, high in crude protein content, high in flavor amino acid content, delicious in taste, more popular with consumers and good in market prospect.

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.