Method for constructing a specialized line of litopenaeus vannamei nitrite tolerance trait

Abstract

The application discloses a method for constructing a dynamic specialized line of Litopenaeus vannamei nitrite tolerance traits. The method comprises the following steps: collecting and cultivating a nitrite tolerance population, screening the nitrite tolerance population, screening three rounds of individuals in the nitrite tolerance population, cultivating zero-generation parents and breeding F1 generation, cultivating the F1 generation and breeding F2 generation, cultivating the F2 generation and breeding F3 generation, and dynamically maintaining the nitrite tolerance trait specialized line. The specialized line breeding method of the Litopenaeus vannamei has the characteristics of simplicity, high efficiency and low cost, and does not need to construct a family line, thereby laying a technical foundation for large-scale and extensive breeding of the Litopenaeus vannamei nitrite tolerance traits.

Description

Technical Field

[0001] This invention belongs to the field of aquaculture technology, specifically relating to a method for constructing a specialized strain of Litopenaeus vannamei with nitrite tolerance. Background Technology

[0002] Nitrite nitrogen and ammonia nitrogen are important pollutants in intensive shrimp farming environments. Currently, nitrite nitrogen has become one of the main stress factors in shrimp farming systems, especially in factory farming and small-scale greenhouse farming models. Excessive nitrite nitrogen levels and difficulty in control have become prominent problems, seriously affecting shrimp growth rate and survival rate. At present, there are no new shrimp varieties with nitrite tolerance on the market. Therefore, it is urgent to cultivate new shrimp varieties with outstanding nitrite tolerance.

[0003] Currently, my country has 15 new varieties of Litopenaeus vannamei, but these new varieties are mainly obtained through family pedigree construction, which requires building a large number of families generation by generation during the breeding process. The construction of shrimp families, the selection of offspring families, and mating require a significant amount of manpower and space in aquaculture facilities, resulting in enormous costs for breeding new shrimp varieties and limiting the enthusiasm of enterprises to participate in this field. Furthermore, to avoid inbreeding, pedigree propagation generally involves mating between different families. This leads to increasingly complex pedigrees in offspring and may also result in the loss of homozygous traits and genotypes. Therefore, it is necessary to explore low-cost and efficient methods for breeding new Litopenaeus vannamei varieties.

[0004] In the production of commercial pigs and other livestock, specialized strains with prominent traits are usually bred, and then combined with strict combining ability tests are conducted to form special hybrid combinations to produce high-quality commercial pigs. This allows the commercial pigs to combine the advantages of each strain and have more obvious hybrid vigor. Shrimp are lower invertebrates and have characteristics that are significantly different from livestock in terms of reproductive biology. Specifically, (1) breeding shrimp can only produce for one breeding season, while excellent breeding stock of livestock such as pigs can produce for many consecutive years; (2) shrimp population reproduction follows an R strategy, where a single adult can produce millions of offspring, but only a few offspring have the opportunity to reproduce in the natural environment. In contrast, the reproduction of higher animal populations such as livestock follows a K strategy, where a single adult can only produce a few high-quality offspring, maintaining population reproduction with a low mortality rate. These two characteristics determine that the specialized strain breeding methods for livestock such as pigs are not suitable for specialized strain breeding of shrimp. Therefore, it is necessary to develop unique specialized strain breeding methods based on the biological characteristics and breeding trait requirements of shrimp. Summary of the Invention

[0005] The purpose of this invention is to provide a method for constructing specialized strains of Litopenaeus vannamei with nitrite tolerance that does not require family construction, and is simple, efficient, and low-cost, thus laying a technical foundation for large-scale and widespread breeding of shrimp with nitrite tolerance.

[0006] To achieve the above objectives, the present invention adopts the following technical solution:

[0007] A method for constructing a specialized strain of Litopenaeus vannamei with nitrite tolerance includes the following steps:

[0008] (1) Collection and cultivation of nitrite-tolerant populations: Collect 10,000 to 20,000 shrimp larvae from 20 to 40 different sources, with a body length of 2 to 3 cm, and place them in the same size preservation pond in the preservation workshop;

[0009] (2) Nitrite-tolerant population screening: 200 shrimp larvae were extracted from each of the above populations and temporarily held in ordinary seawater in the test tank for 48 hours. Sodium nitrite was added to make the final NO2-N concentration 100mg / L (as nitrogen). They were then fed according to conventional feeding and management methods. After 96 hours of stress resistance screening, the activity and feeding of shrimp in the pond were observed 4 times a day. Dead shrimp were removed in time. After 96 hours, the number of surviving shrimp was counted and the survival rate was calculated. The 10-20 populations with the highest survival rate were retained, and the rest of the populations were eliminated.

[0010] (3) First round of screening of individuals in nitrite-tolerant populations: 10,000 shrimp larvae were randomly selected from each population, with an average body length of 2-3 cm, and temporarily held in ordinary seawater for 48 hours; sodium nitrite was added to the clean seawater in the screening pond to make the final NO2-N concentration 100 mg / L; the test shrimp larvae were transferred to the screening pond to start the stress-resistant screening; they were fed according to the conventional feeding and management methods; after 96 hours of stress-resistant screening, the activity and feeding of shrimp in the pond were observed 4 times a day, and shrimp that were sluggish, had empty intestines and stomachs, were dead or dying were removed in time. The shrimp that survived after 96 hours were the shrimp breeding materials that passed the first test;

[0011] (4) Second round of screening of individuals in the nitrite-tolerant population: After 60 days of indoor cultivation, the second nitrite tolerance screening was carried out according to the test method of step (3) above. The stress screening time was 96 hours. The feeding status of shrimp in the pond was observed in the morning, noon and evening every day. Shrimp that were sluggish, had empty intestines and stomachs, died or were dying were removed in time. The shrimp that survived after 96 hours were the breeding materials that passed the second screening and continued the normal cultivation process.

[0012] (5) Third round of screening of individuals in nitrite-tolerant populations: After 90 days of indoor cultivation, the third stress-resistant screening was carried out according to the testing method in step (3) above; after 96 hours of stress-resistant screening, the shrimp were fed normally, and the feeding situation of the shrimp in the screening pond was observed in the morning, noon and evening every day. Shrimp that were sluggish, had empty intestines and stomachs, died or were dying were removed in time, and small shrimp were removed; after 96 hours, the shrimp that survived were the breeding materials that passed the third screening. The surviving individuals in each group were temporarily raised for 3 days. Large, healthy, and normally feeding shrimp individuals were selected from 10 to 20 groups, totaling 2,000 to 3,000 individuals, and the groups were mixed as the zero generation and continued to be cultivated in the parent culture pond;

[0013] (6) Zero-generation parent stock cultivation and F1 generation breeding: After the above 2,000-3,000 nitrite-tolerant shrimp are cultivated to 10-12 months of age, the males and females are separated and raised in separate parent stock ponds at a density of 400-600 shrimp / pond. The parents are then subjected to routine broodstock maturation and management. Mature female shrimp are promptly removed and placed in the male shrimp pond for natural mating. After mating, the female shrimp are removed and placed in spawning tanks for routine spawning and hatching. The offspring of all female shrimp from the same batch are collected in the same nursery pond for routine shrimp seedling cultivation to produce F1 generation closed-group shrimp seedlings with fast growth characteristics.

[0014] (7) F1 generation cultivation and F2 generation breeding: Take 16,000 to 24,000 shrimp F1 generation closed population seedlings (P6 to P10) with nitrite tolerance traits, continue to cultivate to 2 to 3 cm, and conduct the first round of nitrite tolerance trait screening according to the testing method in step (3) above, retaining 8,000 to 12,000 surviving shrimp individuals; continue to cultivate to 60 days, and conduct the second round of nitrite tolerance trait screening according to the testing method in step (3) above, retaining 4,000 to 6,000 surviving shrimp individuals; continue to cultivate to 90 days, and conduct the second round of nitrite tolerance trait screening according to the testing method in step (3) above, retaining 2,000 to 3,000 surviving shrimp individuals, and carry out F1 generation parent cultivation according to the method in step (6) above, and carry out F2 generation breeding according to the method in step (6) above;

[0015] (8) F2 generation cultivation and F3 generation breeding: Take 16,000 to 24,000 shrimp F2 generation closed population seedlings (P6 to P10) with nitrite tolerance traits, continue to cultivate to 2 to 3 cm, and conduct the first round of nitrite tolerance trait screening according to the testing method in step (3) above, retaining 8,000 to 12,000 surviving shrimp individuals; continue to cultivate to 60 days, and conduct the second round of nitrite tolerance trait screening according to the testing method in step (3) above, retaining 4,000 to 6,000 surviving shrimp individuals; continue to cultivate to 90 days, and conduct the second round of nitrite tolerance trait screening according to the testing method in step (3) above, retaining 2,000 to 3,000 surviving shrimp individuals, and carry out F2 generation parent cultivation according to the method in step (6) above, and carry out F3 generation breeding according to the method in step (6) above. The F3 generation is the obtained specialized strain of Litopenaeus vannamei with nitrite tolerance traits;

[0016] (9) Dynamic maintenance of specialized strains with nitrite tolerance: Starting from the F3 generation, 2 to 4 shrimp populations with nitrite tolerance from different sources are reconstructed according to the methods in steps (1) to (5) above. 75 to 100 nitrite-tolerant shrimp individuals are taken from each population, totaling 200 to 300 individuals. These individuals are then raised to the broodstock and added to the specialized strains containing 1,800 to 2,700 broodstock shrimp. Conventional breeding operations are then carried out. This step is repeated once every two generations to complete the dynamic maintenance of specialized strains with nitrite tolerance.

[0017] In step (3), the total volume of the clean seawater in the screening tank is 3m³. 3 .

[0018] In step (5), the weight of the zero-generation shrimp is ≥15g.

[0019] In step (6), the weight of the shrimp when separating males and females is ≥40g.

[0020] This invention also provides the application of the above-described construction method in the breeding of Litopenaeus vannamei. The breeding involves constructing specialized Litopenaeus vannamei strains with nitrite tolerance.

[0021] Compared with traditional family construction methods, the method for constructing specialized shrimp strains with nitrite tolerance of the present invention has the following advantages and positive effects:

[0022] (1) The process of constructing specialized shrimp strains with nitrite tolerance in this invention does not require any family construction operations, which greatly reduces labor intensity and saves a lot of manpower and space for breeding facilities. Therefore, it makes shrimp breeding work simple and easy, laying a technical foundation for the large-scale and widespread breeding of shrimp with fast growth traits.

[0023] (2) In the past, in order to avoid inbreeding, it was necessary to establish families from different sources and carry out complicated inter-family pairing operations and pedigree records. In this invention, since individuals with nitrite tolerance traits that have been strictly screened from 10 to 20 shrimp populations from different sources are mixed in equal proportions, theoretically the maximum probability of inbreeding is only 5 to 10%. Even if there are undesirable offspring from inbreeding, they will be eliminated in the three rounds of screening of breeding materials due to their trait disadvantage.

[0024] (3) In conventional breeding methods centered on family selection, the phenotypic performance of test shrimp is usually used to characterize the traits of the source population or family, thereby selecting the family. However, the test shrimp themselves cannot be used as breeding material. In this invention, after rigorous nitrite stress testing, test shrimp individuals with good performance are directly used as breeding material, which can accurately reflect the traits of the breeding material rather than the traits of the family, greatly improving the accuracy of phenotypic selection and the frequency of occurrence of phenotypic related genotypes.

[0025] (4) Due to cost and facility space limitations, the number of parental individuals in each generation of a family is very limited, which leads to a decrease in the genetic diversity of offspring. In contrast, the parental sources in each generation of this invention are very extensive, and while the traits are gradually purified and the genotypes associated with the traits are gradually homozygous, a high level of genetic diversity is maintained.

[0026] (5) Starting from the F3 generation, every two generations, a new population with nitrite tolerance traits is reintroduced to maintain the specialization of strains in a relatively open and dynamic manner, thereby increasing genetic diversity and reducing the occurrence of genetic degradation. Attached Figure Description

[0027] Figure 1 This is a technical route for constructing specialized strains of Litopenaeus vannamei with dynamic nitrite tolerance. Detailed Implementation

[0028] The following embodiments are further illustrations of the present invention, but not limitations thereof.

[0029] Example 1

[0030] From 2020 to 2024, the specialized strain RN-1 for Litopenaeus vannamei nitrite tolerance was constructed and dynamically maintained in the following manner.

[0031] (1) Collection and cultivation of nitrite-tolerant populations: In May 2020, 15,000 Litopenaeus vannamei larvae from 30 different sources were collected, with a body length of 2-3 cm, and placed in the same size preservation pond in the preservation workshop.

[0032] (2) Nitrite-tolerant population screening: 200 shrimp larvae were extracted from each of the above populations and temporarily held in ordinary seawater in a 200L test tank for 48 hours. Sodium nitrite was then added to achieve a final NO2-N concentration of 100mg / L (as nitrogen). The shrimp were then fed according to conventional feeding and management methods. After 96 hours of stress screening, the activity and feeding of the shrimp in the tank were observed four times daily. Dead shrimp were promptly removed. After 96 hours, the number of surviving shrimp was counted, the survival rate was calculated, and the 15 populations with the highest survival rates were retained.

[0033] (3) First round of screening of individuals in nitrite-tolerant populations: In May 2020, 10,000 shrimp larvae were randomly selected from each of the above 15 populations, with an average body length of 2-3 cm, and were temporarily held in ordinary seawater for 48 hours; in an area of ​​10m² 2 The screening tank was filled with 30cm of clean seawater, with a total volume of 3m³. 3 Sodium nitrite was added to bring the final NO2-N concentration to 100 mg / L. The test shrimp larvae were then transferred to a selection pond to begin stress-resistance selection. They were then fed according to standard feeding and management methods. After 96 hours of stress-resistance selection, the activity and feeding status of the shrimp in the pond were observed four times daily. Shrimp that were sluggish, had empty intestines or stomachs, were dead, or were near death were promptly removed. Shrimp that survived after 96 hours were considered breeding material that passed the first test.

[0034] (4) Second round of screening of individuals in nitrite-tolerant population: After 60 days of indoor cultivation, the second screening of nitrite tolerance was carried out according to the test method of step (3) above. The stress screening time was 96 hours. The feeding status of shrimp in the pond was observed in the morning, noon and evening every day. Shrimp that were sluggish, empty intestines and stomachs, dead and dying were removed in time. The shrimp that survived after 96 hours were the breeding materials that passed the second screening and continued the normal cultivation process.

[0035] (5) Third round of screening of individuals in nitrite-tolerant populations: After 90 days of indoor cultivation, the third stress-resistance screening was carried out according to the testing method of step (3) above; after 96 hours of stress-resistance screening, the shrimp were fed normally, and the feeding situation of the shrimp in the pond was observed in the morning, noon and evening every day. Shrimp that were sluggish, had empty intestines and stomachs, died or were dying were removed in time, and small shrimp were removed; after 96 hours, the shrimp that survived were the breeding materials that passed the third screening. The surviving individuals in each group were temporarily raised for 3 days. A total of 2,500 shrimp individuals (weight ≥15g) with large size, healthy appearance and normal feeding were selected from 15 groups and mixed into the groups as the zero generation, and continued to be cultivated in the parent breeding pond.

[0036] (6) Zero-generation broodstock rearing and F1 generation breeding: After the above 2500 nitrite-tolerant shrimp were raised to 11 months of age (weight ≥40g), males and females were separated and cultured in separate broodstock ponds at a rate of 500 shrimp / pond, and routine broodstock maturation and management were carried out. In April 2021, mature female shrimp were promptly removed and placed in the male shrimp pond for natural mating. After mating, the female shrimp were removed and placed in spawning tanks for routine spawning and hatching operations. The offspring of all female shrimp from the same batch were collected in the same nursery pond for routine shrimp larvae rearing operations, producing F1 generation closed-group shrimp larvae with fast growth traits.

[0037] (7) F1 generation cultivation and F2 generation breeding: In May 2021, 20,000 shrimp F1 generation closed population seedlings with nitrite tolerance traits (P8) were taken and continued to be cultivated to 2-3cm. The first round of nitrite tolerance trait screening was carried out according to the testing method of step (3) above, and 10,000 shrimp individuals were retained. The shrimp were cultivated for 60 days and the second round of nitrite tolerance trait screening was carried out according to the testing method of step (3) above, and 5,000 shrimp individuals were retained. The shrimp were cultivated for 90 days and the second round of nitrite tolerance trait screening was carried out according to the testing method of step (3) above, and 2,500 shrimp individuals were retained. The F1 generation parent stock was cultivated according to the method in step (6) above. In April 2022, the F2 generation was bred according to the method in step (6) above.

[0038] (8) F2 generation cultivation and F3 generation breeding: In May 2022, 20,000 shrimp F2 generation closed population seedlings (P8) with nitrite tolerance were taken and cultivated to 2-3 cm. The first round of nitrite tolerance screening was carried out according to the testing method in step (3) above, and 10,000 shrimp individuals were retained. The shrimp were cultivated for 60 days, and the second round of nitrite tolerance screening was carried out according to the testing method in step (3) above, and 5,000 shrimp individuals were retained. The shrimp were cultivated for 90 days, and the second round of nitrite tolerance screening was carried out according to the testing method in step (3) above, and 2,500 shrimp individuals were retained. The F2 generation parent stock was cultivated according to the method in step (6) above. In April 2023, the F3 generation was bred according to the method in step (6) above. The F3 generation is the obtained Litopenaeus vannamei specialized strain RN-1 with nitrite tolerance.

[0039] (2) Dynamic maintenance of specialized strains with nitrite tolerance: In May 2023, starting from the F3 generation, three shrimp populations with nitrite tolerance from different sources were reconstructed according to steps (1) to (5) above. 83, 83, and 84 nitrite-tolerant shrimp individuals (a total of 250) were taken from each population and raised to parent stock. These were then added to the specialized strain containing 2250 parent shrimp and subjected to routine breeding operations. This step was repeated once every two generations thereafter to maintain the specialized strains with nitrite tolerance.

[0040] Example 2

[0041] From 2021 to 2024, the specialized strain RN-2 for Litopenaeus vannamei nitrite tolerance was constructed using the following method.

[0042] (1) Collection and cultivation of nitrite-tolerant populations: In May 2021, 20,000 Litopenaeus vannamei larvae from 40 different sources were collected, with a body length of 2-3 cm, and placed in the same size preservation pond in the preservation workshop.

[0043] (2) Nitrite-tolerant population screening: 200 shrimp larvae were extracted from each of the above populations and temporarily held in ordinary seawater in a 200L test tank for 48 hours. Sodium nitrite was then added to achieve a final NO2-N concentration of 100mg / L (as nitrogen). The shrimp were then fed according to conventional feeding and management methods. After 96 hours of stress resistance screening, the activity and feeding of the shrimp in the tank were observed four times daily. Dead shrimp were promptly removed. After 96 hours, the number of surviving shrimp was counted, the survival rate was calculated, and the 20 populations with the highest survival rates were retained.

[0044] (3) First round of screening of individuals in nitrite-tolerant populations: In May 2021, 10,000 shrimp larvae were randomly selected from each of the above 20 populations, with an average body length of 2-3 cm, and temporarily held in ordinary seawater for 48 hours; in an area of ​​10m² 2 The screening tank was filled with 30cm of clean seawater, with a total volume of 3m³. 3 Sodium nitrite was added to bring the final NO2-N concentration to 100 mg / L. The test shrimp larvae were then transferred to a selection pond to begin stress-resistance selection. They were then fed according to standard feeding and management methods. After 96 hours of stress-resistance selection, the activity and feeding status of the shrimp in the pond were observed four times daily. Shrimp that were sluggish, had empty intestines or stomachs, were dead, or were near death were promptly removed. Shrimp that survived after 96 hours were considered breeding material that passed the first test.

[0045] (4) Second round of screening of individuals in nitrite-tolerant population: After 60 days of indoor cultivation, the second screening of nitrite tolerance was carried out according to the test method of step (3) above. The stress screening time was 96 hours. The feeding status of shrimp in the pond was observed in the morning, noon and evening every day. Shrimp that were sluggish, empty intestines and stomachs, dead and dying were removed in time. The shrimp that survived after 96 hours were the breeding materials that passed the second screening and continued the normal cultivation process.

[0046] (5) Third round of screening of individuals in nitrite-tolerant populations: After 90 days of indoor cultivation, the third stress-resistance screening was carried out according to the testing method of step (3) above; after 96 hours of stress-resistance screening, the shrimp were fed normally, and the feeding situation of the shrimp in the pond was observed in the morning, noon and evening every day. Shrimp that were sluggish, empty intestines and stomachs, dead and dying were removed in time, and small shrimp were removed; after 96 hours, the shrimp that survived were the breeding materials that passed the third screening. The surviving individuals in each group were temporarily raised for 3 days. Large, healthy, and normally feeding shrimp individuals (weight ≥15g) were selected from 20 groups, totaling 3000 individuals, and were mixed into the groups as the zero generation, and continued to be cultivated in the parent breeding pond.

[0047] (6) Zero-generation broodstock rearing and F1 generation breeding: After the above 3000 nitrite-tolerant shrimp were raised to 11 months of age (weight ≥40g), males and females were separated and cultured in separate broodstock ponds at a rate of 600 shrimp / pond, and routine broodstock maturation and management were carried out. In April 2022, mature female shrimp were promptly removed and placed in the male shrimp pond for natural mating. After mating, the female shrimp were removed and placed in spawning tanks for routine spawning and hatching operations. The offspring of all female shrimp from the same batch were collected in the same nursery pond for routine shrimp larvae rearing operations, producing F1 generation closed-group shrimp larvae with fast growth traits.

[0048] (7) F1 generation cultivation and F2 generation breeding: In May 2022, 24,000 shrimp F1 generation closed population seedlings (P10) with nitrite tolerance were taken and cultivated to 2-3 cm. The first round of nitrite tolerance screening was carried out according to the testing method in step (3) above, and 12,000 shrimp individuals were retained. The shrimp were cultivated for 60 days and the second round of nitrite tolerance screening was carried out according to the testing method in step (3) above, and 6,000 shrimp individuals were retained. The shrimp were cultivated for 90 days and the second round of nitrite tolerance screening was carried out according to the testing method in step (3) above, and 3,000 shrimp individuals were retained. The F1 generation parent stock was cultivated according to the method in step (6) above. In April 2023, the F2 generation was bred according to the method in step (6) above.

[0049] (8) F2 generation cultivation and F3 generation breeding: In May 2023, 24,000 shrimp fry (P10) of the closed population of F2 generation with nitrite tolerance were taken and cultivated to 2-3 cm. The first round of nitrite tolerance screening was carried out according to the testing method in step (3) above, and 12,000 shrimp individuals were retained. The shrimp were cultivated for 60 days and the second round of nitrite tolerance screening was carried out according to the testing method in step (3) above, and 6,000 shrimp individuals were retained. The shrimp were cultivated for 90 days and the second round of nitrite tolerance screening was carried out according to the testing method in step (3) above, and 3,000 shrimp individuals were retained. The F2 generation parent stock was cultivated according to the method in step (6) above. In April 2024, the F3 generation was bred according to the method in step (6) above. The F3 generation is the obtained specialized strain of Litopenaeus vannamei RN-2 with nitrite tolerance.

[0050] Example 3

[0051] In June 2024, 100 larvae of the nitrite-tolerant Litopenaeus vannamei specialized strain RN-2 and 100 larvae of the commercially available ZD strain were collected. Both groups of larvae had a body length of 2-3 cm. Each group of 100 larvae was placed in a test tank containing 100L of seawater with a final NO2-N concentration of 100 mg / L. The test lasted for 96 hours. The activity and feeding of the shrimp in the tank were observed four times daily, and dead shrimp were promptly removed. After 96 hours, the number of surviving shrimp was counted, and the survival rate was calculated. The results showed that the survival rate of the specialized strain RN-2 was 56%, while the survival rate of the ZD strain was 44%. The nitrite-tolerant survival rate of the Litopenaeus vannamei specialized strain RN-2 was 27.3% higher than that of the conventional ZD strain.

[0052] The above are merely preferred embodiments of the present invention. It should be noted that the above preferred embodiments should not be considered as limitations on the present invention, and the scope of protection of the present invention should be determined by the scope defined in the claims. For those skilled in the art, several improvements and modifications can be made without departing from the spirit and scope of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for constructing a specialized strain of Litopenaeus vannamei with nitrite tolerance, characterized in that, Includes the following steps: (1) Collection and cultivation of nitrite-tolerant populations: Collect 10,000 to 20,000 shrimp larvae from 20 to 40 different sources, with a body length of 2 to 3 cm, and place them in the same size preservation pond in the preservation workshop; (2) Nitrite-tolerant population screening: 200 shrimp larvae were extracted from each of the above populations and temporarily held in ordinary seawater in the test tank for 48 hours. Sodium nitrite was added to make the final NO2-N concentration 100 mg / L and the shrimp were fed according to conventional feeding and management methods. After 96 hours of stress screening, the activity and feeding of shrimp in the pond were observed 4 times a day. Dead shrimp were removed in time. After 96 hours, the number of surviving shrimp was counted and the survival rate was calculated. The 10-20 populations with the highest survival rate were retained and the rest were eliminated. (3) First round of screening of individuals in nitrite-tolerant populations: 10,000 shrimp larvae were randomly selected from each population, with an average body length of 2-3 cm, and temporarily held in ordinary seawater for 48 hours; sodium nitrite was added to the clean seawater in the screening pond to make the final NO2-N concentration 100 mg / L; the test shrimp larvae were transferred to the screening pond to start the stress-resistant screening; they were fed according to the conventional feeding and management methods; after 96 hours of stress-resistant screening, the activity and feeding of shrimp in the pond were observed 4 times a day, and shrimp that were sluggish, empty intestines and stomachs, dead or dying were removed in time. The shrimp that survived after 96 hours were the shrimp breeding materials that passed the first test. (4) Second round of screening of individuals in the nitrite-tolerant population: After 60 days of indoor cultivation, the second nitrite tolerance screening was carried out according to the test method of step (3) above. The stress screening time was 96 hours. The feeding status of shrimp in the pond was observed in the morning, noon and evening every day. Shrimp that were sluggish, had empty intestines and stomachs, died or were dying were removed in time. The shrimp that survived after 96 hours were the breeding materials that passed the second screening and continued the normal cultivation process. (5) Third round of screening of individuals in nitrite-tolerant populations: After 90 days of indoor cultivation, the third stress-resistant screening was carried out according to the testing method of step (3) above; after 96 hours of stress-resistant screening, the shrimp were fed normally, and the feeding situation of the shrimp in the screening pond was observed in the morning, noon and evening every day. Shrimp that were sluggish, had empty intestines and stomachs, died or were dying were removed in time, and small shrimp were removed; after 96 hours, the shrimp that survived were the breeding materials that passed the third screening. The surviving individuals in each group were temporarily raised for 3 days. Large, healthy, and normally feeding shrimp individuals were selected from 10 to 20 groups, totaling 2,000 to 3,000 individuals, and the groups were mixed as the zero generation and continued to be cultivated in the parent breeding pond; (6) Zero-generation parent stock cultivation and F1 generation breeding: After the above 2000-3000 nitrite-tolerant shrimp are cultivated to 10-12 months of age, the males and females are separated and raised in parent stock ponds at a density of 400-600 shrimp / pond. The parents are then subjected to routine broodstock maturation and management. Mature female shrimp are promptly removed and placed in male shrimp ponds for natural mating. After mating, the female shrimp are removed and placed in spawning tanks for routine spawning and hatching. The offspring of all female shrimp from the same batch are collected in the same nursery pond for routine shrimp seedling cultivation to produce closed-group F1 generation shrimp seedlings with nitrite tolerance. (7) F1 generation cultivation and F2 generation breeding: Take 16,000 to 24,000 shrimp F1 generation closed population seedlings with nitrite tolerance traits, continue to cultivate to 2 to 3 cm, and conduct the first round of nitrite tolerance trait screening according to the testing method in step (3) above, retaining 8,000 to 12,000 surviving shrimp individuals; continue to cultivate to 60 days, conduct the second round of nitrite tolerance trait screening according to the testing method in step (3) above, retaining 4,000 to 6,000 surviving shrimp individuals; continue to cultivate to 90 days, conduct the third round of nitrite tolerance trait screening according to the testing method in step (3) above, retaining 2,000 to 3,000 surviving shrimp individuals, carry out F1 generation parent breeding according to the method in step (6) above, and carry out F2 generation breeding according to the method in step (6) above; (8) F2 generation cultivation and F3 generation breeding: Take 16,000 to 24,000 shrimp F2 generation closed population seedlings with nitrite tolerance traits, continue to cultivate to 2 to 3 cm, and conduct the first round of nitrite tolerance trait screening according to the testing method of step (3) above, retaining 8,000 to 12,000 surviving shrimp individuals; continue to cultivate to 60 days, conduct the second round of nitrite tolerance trait screening according to the testing method of step (3) above, retaining 4,000 to 6,000 surviving shrimp individuals; continue to cultivate to 90 days, conduct the third round of nitrite tolerance trait screening according to the testing method of step (3) above, retaining 2,000 to 3,000 surviving shrimp individuals, carry out F2 generation parent breeding according to the method in step (6) above, and carry out F3 generation breeding according to the method in step (6) above. The F3 generation is the obtained specialized strain of Litopenaeus vannamei with nitrite tolerance traits; (9) Dynamic maintenance of specialized strains with nitrite tolerance: Starting from the F3 generation, 2 to 4 shrimp populations with different sources with nitrite tolerance were reconstructed according to the methods in steps (1) to (5) above. 75 to 100 nitrite-tolerant shrimp individuals were taken from each population, totaling 200 to 300 individuals. These individuals were then raised to parent stock and added to the specialized strains containing 1,800 to 2,700 nitrite-tolerant shrimp individuals. Conventional breeding operations were then carried out. After that, this step was repeated once every two generations to complete the dynamic maintenance of specialized strains with nitrite tolerance.

2. The method for constructing a specialized strain of Litopenaeus vannamei with nitrite tolerance according to claim 1, characterized in that, In step (3), the total volume of the clean seawater in the screening tank is 3m³. 3 .

3. The method for constructing a specialized strain of Litopenaeus vannamei with nitrite tolerance according to claim 1, characterized in that, In step (5), the weight of the parent shrimp of the zero generation is ≥15g.

4. The method for constructing a specialized strain of Litopenaeus vannamei with nitrite tolerance according to claim 1, characterized in that, In step (6), the weight of the shrimp when separating males and females is ≥40g.

5. The application of the construction method according to any one of claims 1-4 in the breeding of Litopenaeus vannamei.

6. The application according to claim 5, characterized in that, The breeding program aims to develop specialized strains of Litopenaeus vannamei with nitrite tolerance.

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