Method for creating fertile heterologous heptaploid prussian carp gynogenetic line capable of resisting herpesvirus of prussian carp

Abstract

The invention belongs to the technical field of fish genetic breeding, and particularly relates to a creation method of a crucian carp herpesvirus resistant fertile heterologous heptaploid crucian carp gynogenetic germline. The method comprises the following steps: firstly, breeding a gynogenesis double triploid prussian carp female parent and a sexual reproduction diploid megalobrama amblycephala male parent to obtain a filial generation G1, and then screening an individual of which the genome is fused with a set of megalobrama amblycephala sperm chromosome group from the filial generation G1, namely the heterologous heptaploid prussian carp which has the gynogenesis capacity. Cassius auratus gibelio herpesvirus infection experiments prove that the heterologous heptaploid Cassius auratus gibelio clone line has 100% anti-herpesvirus ability, and the breeding route provided by the invention can create a new Cassius auratus gibelio line with significant anti-herpesvirus ability and maintenance of gynogenesis ability, is an important way for breeding new Cassius auratus gibelio varieties, and has wide application prospects.

Description

Technical Field

[0001] The present invention belongs to the technical field of fish genetic breeding, and particularly relates to a method for creating a fertile gynogenetic line of allotetraploid silver crucian carp resistant to carp herpesvirus. Background Art

[0002] As a classical and effective cell engineering breeding technology, polyploid breeding has been widely applied in the breeding of animals and plants, including the variety selection of cultured fish and shellfish. Allotetraploids are produced by the integration and doubling of two different parental genomes, and they often combine the advantages of hybridization and polyploidization, showing better growth advantages and stronger stress resistance than their diploid ancestors. Optimizing or developing new and efficient ploidy manipulation technologies to create excellent germplasms and provide core breeding materials for new variety cultivation is one of the important directions in the genetic breeding of aquaculture animals.

[0003] Allogynogenetic silver crucian carp ( C. gibelio ) is an important aquaculture species in China. However, due to the widespread high-density and single-culture mode in the main silver crucian carp aquaculture areas, an epidemic disease caused by carp herpesvirus ( Ca HV) has broken out in the main silver crucian carp aquaculture areas and caused huge economic losses. Among them, the main cultured variety "Zhongke No. 3" in the crucian carp aquaculture market, that is, the A + line, is extremely susceptible to Ca HV, and the mortality rate after infection reaches 100%. Therefore, synthesizing allogynogenetic silver crucian carp with high resistance to Ca HV is an important requirement for the crucian carp aquaculture industry.

[0004] Silver crucian carp has multiple reproductive modes such as parthenogenetic gynogenesis and sexual reproduction. When performing allospermia gynogenesis, silver crucian carp also has the ability to incorporate chromosome sets and chromosome fragments of sperm from other fish into the egg nucleus for coordinated development. Using this ability, the applicant team used white crucian carp, red crucian carp, etc. as parents to create a series of new polyploid silver crucian carp. However, the white crucian carp is amphidiploid (AABB), and the synthesized new polyploid silver crucian carp has a set of 50 chromosome sets (AB) from the father, forming a double tetraploid crucian carp with a genome composition of AAABBBB. This excessive ploidy will lead to an imbalance in the nucleus-cytoplasm ratio of the cell, bringing some negative effects, and cannot show the advantages of polyploidy. For example, growth and herpes virus infection experiments on the obtained new polyploid silver crucian carp showed that it had no growth advantage compared with the maternal population and had no significant improvement in its ability to resist herpes virus. The applicant team's previous patent CN115486412B disclosed a method for obtaining a new double triploid clone line by backcrossing the red crucian carp with a new double tetraploid male fish that integrates the chromosome set of the red crucian carp. However, this method diluted the genome composition of the original mother "Zhongke No. 3" in the new polyploid silver crucian carp after two generations of integration and hybridization. Its original excellent traits, such as the silver-black body color of "Zhongke No. 3" that is very popular in the market, will be affected by the red crucian carp genome.

[0005] Therefore, breeding a new polyploid silver crucian carp germplasm with high resistance to crucian carp herpes virus, reduced negative impacts, and better traits remains a technical problem that needs to be solved urgently. Summary of the invention

[0006] In order to solve the problems existing in the prior art, the present invention provides a method for creating a fertile allogeneic heptaploid silver crucian carp gynogenetic reproductive system resistant to crucian carp herpes virus.

[0007] The present invention adopts the following technical solutions: A method for creating a fertile allo-heptaploid silver crucian carp gynogenetic reproductive system resistant to crucian carp herpes virus comprises the following steps: S1. The female parent of gynogenetic triploid silver carp (AAABBB) and the male parent of sexual reproduction diploid bighead carp (CC) were bred to obtain the offspring G1; S2. Individuals (AAABBBC) whose genomes have been integrated with a set of sperm chromosomes of Megalobrama amblycephala were screened from the progeny G1, namely, the alloheptaploid silver carp, which has fertile gynogenetic ability.

[0008] Preferably, the female parent of the gynogenetic triploid silver crucian carp in step S1 is the allogeneic silver crucian carp "Zhongke No. 3".

[0009] Preferably, it also includes the S3 propagation step, using the allogeneic heptaploid silver crucian carp as the female parent, using the sperm of the male parent Xingguo red carp to stimulate the eggs of the female parent, and performing gynogenesis to obtain the offspring G2, which is the allogeneic heptaploid silver crucian carp clone system.

[0010] Preferably, female individuals of the allogynogenetic heptaploid crucian carp are selected for egg extraction inspection, and female fish with good egg maturity are selected for breeding. Individuals with a fertilized egg hatching rate exceeding 20% are allogynogenetic heptaploid crucian carp with fertile gynogenetic ability.

[0011] Preferably, in step S1, the specific breeding method is as follows: during the breeding season of crucian carp, the semen of the male parent is mixed with trypsin solution at a volume ratio of semen:trypsin solution of 1:10, and the semen is treated at 23 °C for 15 min; the eggs of the female parent are taken and mixed with the treated semen for fertilization, and the obtained fertilized eggs continue to hatch; the mass concentration of the trypsin solution is 1%.

[0012] Preferably, in step S1, the method for screening allogynogenetic heptaploid crucian carp from the offspring G1 is as follows: a small amount of blood is taken by cutting the fin rays of the fry, and after adding CyStain DNA 1Step solution and mixing evenly, the DNA content is detected by a flow cytometer. Using the maternal double triploid crucian carp and the paternal blunt snout bream as controls, individuals with a DNA content between the sum of the DNA contents of the double triploid crucian carp and the double triploid crucian carp + blunt snout bream are selected, which are allogynogenetic heptaploid crucian carp.

[0013] The beneficial effects of the present invention are as follows: Since double diploid crucian carps such as white crucian carp and red crucian carp all belong to the genus Carassius, the crucian carp herpesvirus will replicate in their bodies after infection, and there are only differences in the survival rate after infection. Therefore, the allogynogenetic new polyploid crucian carp obtained using white crucian carp, red crucian carp, etc. still has susceptibility, and these double diploid crucian carps are not the best male parents for obtaining silver crucian carp new polyploids with absolute disease resistance advantages. The blunt snout bream belongs to the diploid Cyprinidae fish with a relatively distant genetic relationship with silver crucian carp, and has important characteristics such as fast growth, high back, and similar body color to silver crucian carp. In this application, the inventor team has first proved that the blunt snout bream is not infected with the crucian carp herpesvirus. Therefore, by introducing a set of chromosomes of the blunt snout bream that is not infected with Ca HV into the HV-susceptible "Zhongke No. 3", the new polyploid silver crucian carp obtained can improve the susceptibility of the original female parent to the virus and greatly improve its antiviral ability, thereby obtaining a new polyploid silver crucian carp with absolute disease resistance advantages. Ca In the traditional cross-breeding process, due to the reproductive isolation between the blunt snout bream and the crucian carp, sexual reproduction cannot be carried out to produce fertile hybrid offspring. However, this method utilizes the special reproductive characteristics of silver crucian carp to break through the reproductive isolation between silver crucian carp and blunt snout bream, and obtains fertile allogynogenetic heptaploid silver crucian carp offspring. At the same time, this polyploid silver crucian carp maintains its ability of parthenogenetic gynogenesis, and can permanently fix its disease resistance advantages. The method of this application not only obtains the crucian carp-bream hybrids that are difficult to obtain by traditional hybridization, but also breaks through the traditional understanding that aneuploids are sterile. The herpesvirus infection experiment confirms that the allogynogenetic heptaploid silver crucian carp clone line is infected with

[0014] Ca ​Only trace amounts of virus particles were detected in the body after HV injection, indicating that virus replication in the body was significantly inhibited and thus no disease or death occurred, that is, it had 100% anti-herpes virus ability.

[0015] This application provides an important reference for the selection of parents in polyploid breeding, and species with more distant genetic relationships can be selected for the improvement of more traits. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 It shows the creation route of the allogynogenetic clone line of the allotetraploid silver crucian carp provided by this application.

[0017] Figure 2 It is the histogram of the DNA content of blood cells of the allotetraploid silver crucian carp and its parental stocks in Example 1.

[0018] Figure 3 It is the identification of the genomic composition of the allotetraploid silver crucian carp by genomic in situ hybridization in Example 1. Among them, the blue fluorescence signal is the chromosome stained with DAPI, the green signal is the hybridization signal of the genomic probe of the maternal silver crucian carp labeled with digoxigenin, and the red signal is the hybridization signal of the genomic probe of Megalobrama amblycephala labeled with biotin.

[0019] Figure 4 It is the identification of the genomic composition of the G2 generation silver crucian carp produced by the silver crucian carp screened by genomic in situ hybridization in Example 2. The red fluorescence signal is the chromosome of Megalobrama amblycephala.

[0020] Figure 5 It is the identification of the genomic composition of the non-reproducing G1 generation silver crucian carp by genomic in situ hybridization in Example 2. The red fluorescence signal is the chromosome of Megalobrama amblycephala.

[0021] Figure 6 It shows the survival curves of the allotetraploid silver crucian carp, maternal silver crucian carp and paternal Megalobrama amblycephala after infection with Ca HV.

[0022] Figure 7 It shows the virus copy numbers of the allotetraploid silver crucian carp, maternal silver crucian carp and paternal Megalobrama amblycephala after infection with Ca HV.

[0023] Figure 8 It shows the head kidney tissue pathological sections of the allotetraploid silver crucian carp, maternal silver crucian carp and paternal Megalobrama amblycephala after infection with Ca HV. DETAILED DESCRIPTION OF THE INVENTION

[0024] For ease of understanding, the technical solutions of the present invention will be described in more detail below with reference to specific examples.

[0025] Example 1 Creation of a fertile allogynogenetic clone line of allotetraploid silver crucian carp As Figure 1 shown, the gynogenetic diploid triploid crucian carp female parent (AAABBB) was bred with the sexually reproductive diploid blunt snout bream male parent (CC) to obtain the offspring G1; individuals with DNA content between that of the diploid triploid crucian carp and the sum of the DNA content of the diploid triploid crucian carp and blunt snout bream were screened from the offspring G1, that is, individuals with a set of blunt snout bream sperm chromosome sets incorporated into their genomes (AAABBBC), which are allopolyploid crucian carp, and the allopolyploid crucian carp has fertile gynogenetic ability.

[0026] The specific operations are as follows: 1. Prepare a 1% trypsin solution Weigh 0.1 g of trypsin powder and add it to 10 mL of sperm preservation solution to make its working concentration 1%. Dissolve and mix evenly for standby. Place it at 4°C and use it freshly prepared.

[0027] 2. Treat blunt snout bream sperm with trypsin solution Select sexually mature male blunt snout bream (genome composition is CC, chromosome number is 48) during the breeding season. Through sexual reproductive meiosis, meiotic sperm (C) is produced. Take 1 mL of semen and add it to 10 mL of the above trypsin solution, and treat it at 23°C for 15 min. During this period, invert and mix it repeatedly to make the treatment complete.

[0028] 3. Fertilize the treated sperm with the mature eggs of Zhongke No. 3 Select the allogynogenetic crucian carp "Zhongke No. 3" with mature gonads, inject the ovulation hormone, and put the injected parent fish into a tank with a constant indoor temperature of 21°C and running water. The effective period is about 10 hours. Therefore, the ovulation hormone is injected at night and spawning occurs in the morning for the next experiment. After the allogynogenetic crucian carp "Zhongke No. 3" spawns, squeeze the mature eggs into a 10 cm large dish, and then add the blunt snout bream sperm treated with trypsin for 15 min for dry fertilization. Use a dry feather to stir and mix the semen and eggs evenly and sprinkle them into a white porcelain dish filled with aerated water at 23°C (about 2000 - 4000 fertilized eggs per dish) for cultivation.

[0029] 4. Incubate and select the fertilized eggs During the incubation stage of the fertilized eggs in the white porcelain dish, change the water every 12 h, add a small amount of methylene blue to fix the dead fertilized eggs. After 24 h, pick out the blue dead eggs fixed by methylene blue in the white porcelain dish and remove them to prevent water deterioration and affect the normally developing fertilized eggs. After the surviving fertilized eggs hatch, transfer them to a fish tank for mild aeration and cultivation.

[0030] 5. Ploidy detection and cultivation of candidate offspring When the above fry were cultured to six months old, a trace amount of blood (about 0.1 μL) was collected by cutting the fin rays. The blood was added to 200 μL of 4°C pre-cooled CyStain DNA 1Step (Partec CyStain DNA 1Step, Germany) solution and mixed. The blood was placed on ice and waited for the DNA content to be detected by the machine (CytoFLEX flow cytometer). The female triploid silver carp and the male agglomerate bream were used as controls. The individuals with DNA content between the triploid silver carp and the sum of the triploid silver carp + agglomerate bream were selected, which were allogenic heptaploid silver carp, see Figure 2 .

[0031] In this example, a total of 15 allogeneic heptaploid silver carp (AAABBBC) incorporating a set of chromosome sets from sperm of Megalobrama amblycephala were screened.

[0032] 6. Screening and reproduction of fertile offspring In polyploid breeding, aneuploids are usually infertile due to the inability to undergo normal meiosis, for example, triploid seedless watermelons. In the present application, the identified allogeneic heptaploid silver crucian carp is cultured to sexual maturity, and the gonadal development of the allogeneic heptaploid silver crucian carp is examined, and mature individuals whose oocytes have developed to the fourth stage are selected for reproduction by injecting oxytocin, and their eggs are fertilized with ultraviolet-inactivated Xingguo red carp sperm, and the fertilized eggs are hatched to obtain a batch of allogeneic heptaploid silver crucian carp female nuclear reproductive clone line offspring.

[0033] In the present embodiment, 15 individuals screened were cultured to sexual maturity, and the female fish egg development was checked. Select a female fish parent with a strong body, no deformity and soft abdomen, enlarged, and clear ovarian outline, and use an egg digger to dig eggs for inspection: the egg extractor is slowly inserted into the genital pore, then slightly to the left or right, rotated a few times and gently extracted, a small amount of eggs can be taken out, and then a small amount of fixed transparent liquid is added, and the egg nuclear position is observed after soaking for 2-3 minutes. If the nuclear position of all or most of the eggs is eccentric or polarized, it shows that the egg maturity is good; as the white nucleus occupies the central position, it shows that the egg development is poor; if most of the eggs do not have the white core to appear, they are mostly degenerate eggs or overmature, which is not suitable for breeding. Subsequently, the qualified female fish were bred by injecting oxytocin, and their mature eggs were dry-fertilized with ultraviolet-inactivated Xingguo red carp sperm. After the fertilized eggs hatched out of the membrane, the hatching rate was statistically >20%, and the parents were considered fertile. In this embodiment, one fertile alloheptaploid parent was finally screened, and a batch of gynogenetic clone offspring of this individual were obtained.

[0034] 7. Genetic identification of offspring of gynogenetic clones To verify that all the progenies of the gynogenetic offspring of the allotetraploid gibel carp are heptaploid, 3 progeny individuals were randomly selected to prepare metaphase chromosome spreads. Subsequently, through a whole-genome in situ hybridization experiment, the genome of the female parent gibel carp was labeled with digoxigenin, and the genome of the male parent Megalobrama amblycephala was labeled with biotin. As Figure 3 shown, where Figure a represents the superimposed image of all signals, the red signals in Figure b are the hybridization signals of the probe of the Megalobrama amblycephala genome labeled with biotin, the green signals in Figure c are the hybridization signals of the probe of the gibel carp genome labeled with digoxigenin, and the blue fluorescence signals in Figure d are the chromosomes stained with DAPI. It can be observed that the gynogenetic offspring of the allotetraploid gibel carp are indeed heptaploids with a set of sperm chromosomes of Megalobrama amblycephala (24 chromosomes labeled with red signals) incorporated into the gibel carp.

[0035] Example 2 Method for creating a fertile gynogenetic clone line of allotetraploid gibel carp: In this Example 2, the methods such as fertilization, hatching, cultivation, and ploidy detection of Megalobrama amblycephala and the double-triploid gibel carp are the same as those in Example 1 and will not be elaborated here.

[0036] In this example, blood samples were taken from the obtained G1 individuals for DNA content detection. 13 gibel carps with DNA content between that of the male parent Megalobrama amblycephala (diploid) and the female parent gibel carp (double-triploid) were screened and used as candidate allotetraploid gibel carps for further breeding experiments.

[0037] 10 gibel carps were randomly selected from the 13 screened gibel carps for breeding experiments, and all 10 gibel carps could obtain G2 generations through gynogenesis. From the G2 generations produced by each gibel carp, 3 progeny individuals were randomly selected to prepare metaphase chromosome spreads, and then a whole-genome in situ hybridization experiment was carried out. The detected progenies were all allotetraploid gibel carps. Figure 4 The detection results of 6 randomly selected G2 individuals are shown. Thus, it can be confirmed that all 10 gibel carps that produced G2 generations are fertile allotetraploid gibel carps.

[0038] Chromosome fluorescence in situ hybridization detection was also carried out on the remaining 3 gibel carp individuals that were not screened. The results are shown in Figure 5 , and it can be seen that all 3 gibel carp individuals are also allotetraploid gibel carps.

[0039] Based on the above experiments, it can be confirmed that in the breeding method of this application, the allotetraploid trait and the fertile trait in the G1 generation are highly correlated, and all the obtained allotetraploid gibel carps can perform gynogenesis.

[0040] Example 3 Ca HV artificial infection experiment Prepare fresh Ca HV virus suspension, and use PBS buffer at 10 0, 10 1 , 10 2 , 10 3 , 10 4 The dilution multiples were used for gradient dilution on ice. Sixty healthy Carassius auratus gibelio var. CAS III were randomly divided into six groups (10 fish per group), and were respectively injected with the virus filtrate by gradient dilution. Each fish was injected with 50 μL of the virus filtrate dilution through the base of the pectoral fin, and the control group was injected with an equal amount of PBS buffer. Observation was carried out daily, the mortality was counted, and the median lethal dose was calculated as 10 2 dilution.

[0041] The viral copy number was calculated by fluorescence quantitative PCR for quantification. The helicase gene fragment of HV 637bp was used as the quantification standard to construct a standard curve. The specific steps are as follows: Ca 1) Amplification The 637bp helicase fragment of HV was amplified and purified and recovered using a gel extraction kit. Ca

[0042] 2) The purified and recovered helicase fragment was inserted into the pMD®18-T plasmid to obtain the pMD- Ca HV plasmid (3,329bp).

[0043] 3) The copy number of the plasmid was calculated according to the relationship between the number of moles, relative molecular mass and concentration. The pMD- Ca HV plasmid with known copy number was serially diluted 10-fold as the standard, and was quantitatively analyzed by fluorescence quantitative PCR. The SPSS software was used to construct a standard curve.

[0044] 4) The total DNA of the sample was extracted using a DNA extraction kit, and was used as a template for quantitative analysis by fluorescence quantitative PCR under the same conditions. The viral copy number of each sample was calculated according to the standard curve.

[0045] Ca Finally, the calculated median lethal dose was 50 μL The HV diluted filtrate contained a viral copy number of 3.214×10 7 , and this concentration was used as the injection concentration for subsequent experiments.

[0046] The gynogenetic clone line of allogynogenetic heptaploid Carassius auratus, the maternal double triploid Carassius auratus gibelio var. CAS III, and the paternal Megalobrama amblycephala were temporarily cultured in a 32 cm×24 cm×36 cm aquarium in the laboratory. The breeding water temperature was gradually transitioned to 23°C (±1°C), and the fish were fed for two weeks to observe that there was no abnormality in the experimental fish and PCR was used to detect that there was no Ca HV virus in the fish body. During the temporary culture period, the fish were fed normally every day, and feeding was stopped 24 h before and after virus injection. 50 μL of the Ca virus suspension filtered by HV (the number of virus particles was 3.214×107 ), wild-type individuals were used as the control group, and each was injected with an equal amount of PBS buffer. Three parallel groups were set up for each infection group, and each parallel group had 30 individuals. The culture water was maintained at 23°C (±1°C) and continuously filtered, and changed once a day. When no dead individuals appeared, 1 / 3 of the water was changed daily; after fish diseases occurred and deaths appeared, 1 / 2 of the water was changed daily and the dead individuals were promptly fished out to keep the water clean.

[0047] The death situation of gibel carp was recorded daily after infection until 14 dpi (days post infection), and the survival rate of fish was analyzed. As Figure 6 shown, the results showed that the female parent "Zhongke No. 3" began to die on the 6th day and all died until the 8th day, and the final survival rate was 0%; while the final survival rates of the male parent Megalobrama amblycephala group and the allotriploid gibel carp were both 100%, indicating that the allotriploid gibel carp has a very high ability to resist carp herpesvirus after virus infection.

[0048] Furthermore, the virus copy numbers in the head kidney tissues of the three kinds of fish were calculated. As Figure 7 shown, the female parent "Zhongke No. 3", that is, line A + showed an exponential increase in the virus in the body after being infected with Ca HV virus, while almost no virus particles were detected at different time points after infection in Megalobrama amblycephala, and trace amounts of virus particles could be detected in the allotriploid. At the same time, pathological section analysis of the head kidney tissue was carried out. See Figure 8 , and it was found that 120 h after virus infection, the pathological damage of the head kidney tissue in the allotriploid group was significantly lower than that of the female parent gibel carp line A + .

[0049] These results indicate that the female parent line A + is highly susceptible to Ca HV, the male parent Megalobrama amblycephala is not infected with Ca HV, and although the allotriploid clone line is infected with Ca HV, the virus replication in its body is significantly inhibited and thus it will not get sick and cause death, and it has a very high antiviral ability.

[0050] The above experiments prove that the breeding route provided by this application can create a new gibel carp strain with significant anti-herpesvirus ability and still maintain the ability of gynogenesis, so it is an important way to cultivate new gibel carp varieties.

[0051] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the present invention; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that any modifications, equivalent replacements, and improvements made within the spirit and principle of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for creating a fertile alloheptaploid silver crucian carp gynogenetic reproductive system resistant to crucian carp herpesvirus, characterized in that: The following steps are involved: S1. The female parent of gynogenetic triploid silver carp (AAABBB) and the male parent of sexual reproduction diploid bighead carp (CC) were bred to obtain the offspring G1; S2. Individuals whose genomes have been integrated with a set of sperm chromosomes of Megalobrama amblycephala (AAABBBC) were screened from the progeny G1, namely, the alloheptaploid silver carp, in which the female individuals have fertile gynogenetic ability.

2. The method for creating a fertile alloheptaploid silver carp gynogenetic reproductive system resistant to crucian carp herpesvirus as claimed in claim 1, characterized in that: The female parent of the gynogenetic triploid silver crucian carp in step S1 is the allogynogenetic silver crucian carp "Zhongke No. 3".

3. The method for creating a fertile alloheptaploid silver carp gynogenetic reproductive system resistant to herpesvirus of crucian carp as claimed in claim 1, characterized in that: The method also includes an S3 propagation step, in which a female individual of the allogeneic heptaploid silver crucian carp is used as the female parent, sperm of the male parent Xingguo red carp is used to stimulate the eggs of the female parent, and gynogenesis is performed to obtain offspring G2, which is a clone system of the allogeneic heptaploid silver crucian carp.

4. The method for creating a fertile alloheptaploid silver carp gynogenetic reproductive system resistant to herpesvirus of crucian carp as claimed in claim 3, characterized in that: The eggs of female individuals in the alloheptaploid silver crucian carp were dug out for inspection, and females with well-matured eggs were selected for breeding. Individuals with a fertilized egg hatching rate greater than 20% were alloheptaploid silver crucian carp with fertile female nuclear reproductive ability.

5. The method for creating a fertile alloheptaploid silver carp gynogenetic reproductive system resistant to herpesvirus of crucian carp as claimed in claim 1, characterized in that: In step S1, the specific method of breeding is: in the crucian carp breeding season, the male semen is mixed with the trypsin solution at a volume ratio of 1:10, and the semen is treated at 23°C for 15 minutes; the female eggs are mixed with the treated semen for fertilization, and the obtained fertilized eggs are continued to be hatched; the mass concentration of the trypsin solution is 1%.

6. The method for creating a fertile alloheptaploid silver carp gynogenetic reproductive system resistant to herpesvirus of crucian carp as claimed in claim 1, characterized in that: In the step S1, the method for screening allogeneic heptaploid silver crucian carp from the offspring G1 is as follows: cutting the fin rays of the fry to obtain a trace amount of blood, adding CyStain DNA 1Step solution and mixing, detecting the DNA content by flow cytometer, taking the maternal triploid silver crucian carp and the paternal amblycephala bream as controls, selecting individuals with a DNA content between the triploid silver crucian carp and the sum of the DNA contents of the triploid silver crucian carp + amblycephala bream, that is, the allogeneic heptaploid silver crucian carp.

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