Method for breeding a fast-growing high-survival-rate triploid improved strain of oyster crassostrea hongkongensis

Abstract

The application firstly obtains the shell black and shell gold diploid strains of Fujian oyster with pure shell color and fast growth through population breeding, and then obtains the new diploid Fujian oyster strain with fast growth speed and high survival rate through hybrid breeding. Secondly, the shell black improved strain of tetraploid Fujian oyster with fast growth, high genetic diversity, high tetraploid rate and stability is obtained through population breeding and family breeding. Finally, the fast-growing high-survival-rate improved strain of triploid Fujian oyster is bred by hybridizing the new diploid Fujian oyster strain obtained above as a female parent and the shell black improved strain of tetraploid Fujian oyster as a male parent, which has the excellent characteristics of high survival rate, fast growth speed and high yield while maintaining the high-temperature-resistant characteristics of Fujian oyster, and is suitable for popularization and cultivation in the southern coastal areas of Fujian, Guangdong and other places in China, and has a broad industrial application prospect.

Description

Technical Field

[0001] This invention belongs to the field of shellfish breeding technology, specifically relating to a method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster. Background Technology

[0002] Oyster farming holds an important position in my country's marine aquaculture industry. In recent years, polyploid breeding technology has attracted much attention. Compared with diploid oysters, triploid oysters exhibit advantages such as rapid growth, low reproductive capacity, and large size. Furthermore, triploid oysters, due to their sterility, can still be supplied to the market normally during the summer, meeting consumer demand.

[0003] Since triploids cannot produce viable offspring, genetic improvement of triploids often involves improving diploid or tetraploid parents, and then combining the superior traits of both parents through ploidy breeding and hybridization to achieve the goal of genetic improvement. This method has been widely used in triploid Pacific oysters and triploid American oysters.

[0004] As one of the highest-yielding farmed shellfish, the Fujian oyster is widely cultivated in the southern coastal areas of Fujian and Guangdong provinces in my country. However, diploid Fujian oysters suffer from drawbacks such as small size and long reproductive cycles. In recent years, although triploid Fujian oysters have been widely used industrially, it has been found that traditionally cultivated triploid Fujian oysters exhibit slow growth and high mortality rates. Therefore, there is an urgent need for genetic improvement of traditional triploid Fujian oysters to meet industry demands. Summary of the Invention

[0005] To overcome the problems of slow growth and high mortality rate of traditional triploid Fujian oysters in my country, this invention develops a method for cultivating a fast-growing, high-survival-rate improved triploid Fujian oyster strain, which can provide a theoretical and practical basis for the cultivation and industrialization of superior Fujian oyster varieties.

[0006] A method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster, characterized by the following steps:

[0007] a. Select diploid Fujian oysters with golden and black shells as parent oysters, and improve shell color and growth traits through population selection to obtain diploid Fujian oyster strains with golden and black shells.

[0008] b. By combining the additive genetic variance accumulated within the two strains, Shell Gold and Shell Black, and the non-additive genetic variance between strains during the breeding process through inter-strain hybridization, a new diploid Fujian oyster hybrid strain with fast growth rate and high survival rate can be obtained.

[0009] c. Using the diploid Fujian oyster shell black line as the male and female parents, a triploid Fujian oyster shell black line was induced; using the triploid Fujian oyster shell black line as the female parent and the diploid Fujian oyster shell black line as the male parent, a tetraploid Fujian oyster shell black line was induced.

[0010] d. Through population selection and family selection, the shell color, growth and reproductive capacity of the tetraploid Fujian oyster black-shelled strain are selected in a targeted manner to obtain an improved tetraploid Fujian oyster black-shelled strain with pure shell color, fast growth rate and high reproductive capacity.

[0011] e. Using the diploid Fujian oyster hybrid new line described in step (b) as the female parent and the tetraploid Fujian oyster shell black improved line described in step (d) as the male parent, a fast-growing and high-survival-rate triploid Fujian oyster improved line can be obtained by hybridization.

[0012] Furthermore, in step (a), oysters with good vitality, beautiful and undamaged shells, and pure gold or pure black shells on both sides are preferred as parent oysters for the two basic populations. When selecting individuals for population breeding, the top 10% of individuals in terms of shell height and wet weight are selected and continuously intensively bred for 3 to 4 generations.

[0013] Furthermore, in step (b), when hybridization is carried out, individuals with high shell height and total weight are preferred. At least 50 male and female parents are selected from the black-shelled and gold-shelled strains respectively, and fertilization is performed at a sperm:egg ratio of 50:1.

[0014] Furthermore, in step (c), the triploid Fujian oyster shell black strain is induced by selecting individuals with pure black color, high shell height, and total weight from the diploid Fujian oyster shell black strain as parents. Among them, individuals with uniform development, ≥15 million eggs, and dark yolk color are selected as mothers, and individuals with full gonads and vigorous sperm motility are selected as fathers. Sperm and eggs are obtained by "dry peeling method".

[0015] Furthermore, in step (c), the triploid Fujian oyster shell black strain is induced by fertilization at a sperm:egg ratio of 50:1. After fertilization for 15 minutes, 0.75 mg / L of cytochalasin B is added to inhibit the release of the second polar body of the fertilized egg. After continuous treatment for 15 minutes, the fertilized eggs are soaked in seawater with a 2 mg / L dimethyl sulfoxide solution. After hatching and selection, the ploidy of the larvae is determined, and the group with a triploid rate of more than 95% is retained. The larvae are then raised and the adult oysters are grown using conventional methods to obtain the triploid Fujian oyster shell black induced strain.

[0016] Further, in step (c), the tetraploid Fujian oyster shell black strain is induced by selecting individuals from the triploid Fujian oyster shell black strain that are uniformly developed, have ≥15 million eggs, and have a deep yolk color as the female parent, and selecting individuals from the diploid Fujian oyster shell black strain that have vigorous sperm motility as the male parent. Fertilization is carried out at a sperm:egg ratio of 50:1. 8-10 minutes after fertilization, 1 mg / L of cytochalasin B is added to inhibit the release of the first polar body of the fertilized egg. The group without reagent is used as the control group. When 50% of the fertilized eggs in the control group show the first polar body, the eggs are washed. After hatching and selection, the ploidy of the larvae is determined. The group with a tetraploid rate of more than 80% is retained. The larvae are cultured and the adult oysters are raised using conventional methods to obtain the tetraploid Fujian oyster shell black induced strain.

[0017] Furthermore, in step (d), when conducting group selection, the black-shelled tetraploid Fujian oyster with pure black shell color, high shell height and total weight, and high reproductive capacity is preferred as the parent. The top 10% of individuals in terms of shell height and total weight are selected, and more than 50 males and 50 females are selected in each generation. They are fertilized at a sperm:egg ratio of 50:1, and high-intensity selection is carried out for 2 to 3 generations.

[0018] Furthermore, in step (d), when conducting population selection, step (c) is repeated to obtain black-shelled tetraploid Fujian oysters with different genetic backgrounds, and these are continuously added to the original tetraploid population to ensure the high genetic diversity of the black-shelled tetraploid Fujian oyster population.

[0019] Furthermore, in step (d), when conducting family selection, the best parent is the black-shelled tetraploid Fujian oyster with pure black shell color, high shell height and total weight, and high fertility. 40-50 full-sib families are established through one-to-one fertilization. After the gonads of the previous generation of families mature, the families with pure black shell color and a tetraploid rate of more than 95% are selected as the parents of the next generation of families. One-to-one fertilization is carried out between tetraploid individuals from different families to obtain the next generation of families. After 2-3 generations of family selection, a tetraploid Fujian oyster black-shelled improved strain with high tetraploid rate, pure shell color, and high fertility can be obtained.

[0020] Furthermore, in step (e), individuals with uniform development, ≥15 million eggs, and dark yolk color are selected from diploid Fujian oyster hybrid strains as maternal parents, and individuals with pure black shell color and vigorous sperm motility are selected from tetraploid Fujian oyster shell black improved strains as paternal parents. Fertilization is carried out at a sperm:egg ratio of 50:1, and larval and adult oyster rearing is carried out using conventional methods to obtain a fast-growing and high-survival-rate triploid Fujian oyster improved strain.

[0021] Beneficial effects of the present invention

[0022] The technical solution provided by this invention has the following advantages compared with known technologies.

[0023] (1) Diploid Fujian oysters were genetically improved through selective breeding and hybridization to obtain a new diploid Fujian oyster hybrid strain with fast growth and high survival rate as the female parent. (2) A tetraploid Fujian oyster shell-black improved strain with high and stable tetraploidity, fast growth, and high reproductive capacity was obtained through population selection and family selection to obtain the male parent. (3) While selecting the tetraploid population, repeated induction experiments were conducted to supplement the original population with tetraploids with different genetic backgrounds, further improving the genetic diversity of the tetraploid population. (4) This invention uses a new diploid Fujian oyster hybrid strain as the female parent and a tetraploid Fujian oyster shell-black improved strain with high genetic diversity as the male parent to obtain a fast-growing and high-survival-rate triploid Fujian oyster improved strain. The offspring retain the high-temperature resistance characteristics of Fujian oysters while significantly improving growth rate and survival rate, making them suitable for promotion and cultivation in the southern coastal areas of Fujian and Guangdong in my country, with broad prospects for industrial application. Attached Figure Description

[0024] Figure 1 This is a technical roadmap for the cultivation method of a fast-growing, high-survival-rate triploid improved strain of Fujian oyster according to the present invention. Detailed Implementation

[0025] The present invention will be further illustrated by specific embodiments below, but these are not intended to limit the invention. Example

[0026] a. Five thousand diploid wild populations of *Crassostrea gigas* were collected from Putian, Fujian. Individuals with pure gold and pure black shells were selected as parent oysters. Shell height was chosen as the target trait, and with a selection intensity of 1.5–1.9, 150 golden-shelled and 150 black-shelled oysters with the largest shell height were selected as parent oysters. Fertilization was performed at a sperm:egg ratio of 50:1. Larval and adult rearing were conducted using conventional methods to obtain diploid *Crassostrea gigas* gold-shelled and black-shelled selective breeding lines. Subsequently, the same breeding objectives and techniques were used to conduct three consecutive generations of population selection on the gold-shelled and black-shelled selective breeding lines to obtain the F3 generation of diploid *Crassostrea gigas* gold-shelled and black-shelled selective breeding lines.

[0027] b. After the gonads of the diploid gold-shelled and black-shelled F3 generation oyster breeding lines mature, 100 individuals with pure shell color, shell height and total weight are selected from the two lines as hybrid parents and two new diploid hybrid lines of Fujian oysters are obtained by complete diallel hybridization.

[0028] c. From the diploid Fujian oyster black-shell strain, individuals with pure black shells, high shell height, and large total weight were selected as parents; 30 individuals with uniform development, ≥15 million eggs, and deep yolk color were selected as mothers; and 30 individuals with full gonads and vigorous sperm motility were selected as fathers. Sperm and eggs were obtained using the "dry peeling method." Fertilization was performed at a sperm:egg ratio of 50:1. 15 minutes after fertilization, 0.75 mg / L of cytochalasin B was added to inhibit the release of the second polar body of the fertilized egg. After continuous treatment for 15 minutes, the fertilized eggs were soaked in seawater with a 2 mg / L dimethyl sulfoxide solution. After hatching and selection, the ploidy of the larvae was determined. The group with a triploid rate of over 95% was retained. The larvae were cultured and the adult oysters were raised using conventional methods to obtain the induced triploid Fujian oyster black-shell strain.

[0029] d. From the induced triploid Fujian oyster black-shell strain, select 50 individuals with uniform development, ≥15 million eggs, and deep yolk color as maternal parents, and from the diploid Fujian oyster black-shell strain, select 50 individuals with vigorous sperm motility as paternal parents. Insemination was carried out at a sperm:egg ratio of 50:1. 8-10 minutes after fertilization, 1 mg / L of cytochalasin B was added to inhibit the release of the first polar body of the fertilized egg. The group without reagents was used as the control group. When 50% of the fertilized eggs in the control group showed the first polar body, the eggs were washed. After hatching and selection, the ploidy of the larvae was determined. The group with a tetraploid rate of more than 80% was retained. The larvae were cultured and the adult oysters were raised using conventional methods to obtain the induced tetraploid Fujian oyster black-shell strain.

[0030] e. Select black-shelled tetraploid Fujian oysters with pure black shell color, high shell height and total weight, and high reproductive capacity as parents. Select the top 10% of individuals in terms of shell height and total weight to construct a tetraploid Fujian oyster black-shelled strain. Continuously select and breed in a high-intensity population for 2-3 generations, and select more than 50 males and 50 females in each generation.

[0031] f. Repeat step (e) to obtain black-shelled tetraploid Fujian oysters with different genetic backgrounds, and continuously add them to the original tetraploid population to ensure the high genetic diversity of the black-shelled tetraploid Fujian oyster strain.

[0032] g. Select black-shelled tetraploid Fujian oysters with pure black shell color, high shell height, and high total weight as parents. Establish 40-50 full-sib families through one-to-one fertilization. After the gonads of the previous generation of families mature, select families with pure black shell color and a tetraploid rate of more than 95% as parents for the next generation of families. Randomly fertilize tetraploid individuals from different families one-to-one to obtain the next generation of families. After 2-3 generations of family selection, a tetraploid Fujian oyster black-shelled improved strain with high tetraploid rate, pure shell color, and high reproductive capacity can be obtained.

[0033] h. Using the new diploid Fujian oyster hybrid line described in step (b) as the female parent and the improved tetraploid Fujian oyster black shell line described in step (g) as the male parent, a fast-growing and high-survival-rate triploid Fujian oyster new line can be obtained through hybridization. The obtained triploid Fujian oyster new line, along with the black shell, gold shell, and triploid control groups, were transferred to the main aquaculture sites in Putian, Fujian for trial cultivation. The production data of each group were recorded in Table 1.

[0034] As shown in Table 1, the new fast-growing, high-survival triploid Fujian oyster strain cultivated in this invention has significant production advantages. Compared with the black-shelled and gold-shelled triploid control groups, its growth rate can be increased by 20-40%, its survival rate can be increased by more than 30%, and its yield can be increased by more than 90%. Compared with the black-shelled and gold-shelled diploid control groups, its growth rate can be increased by 104-168%, and its yield can be increased by more than 160%, significantly improving the economic benefits of Fujian oyster farming.

[0035] Table 1. Comparison of production performance between fast-growing, high-survival-rate triploid Fujian oyster strains and the control group.

[0036] Shell height (mm) Wet weight (g) Survival rate (%) Yield (g) New line 88.39 89.65 58.15 156388 Shell black 3n 72.71 63.68 43.21 82548 Shell gold 3n 68.99 63.1 41.89 79291.8 Shell black 2n 43.45 33.5 32.54 32701.7 Shell gold 2n 40.36 32.89 30.82 30414

[0037] The above description is only used to illustrate the technical solutions and features of the present invention, and is not intended to limit the scope of protection of the present invention. Any simple modifications or equivalent substitutions made by those skilled in the art to the technical solutions of the present invention should fall within the scope of protection of the technical solutions of the present invention.

Claims

1. A method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster, characterized in that, Includes the following steps: a. Select diploid Fujian oysters with golden and black shells as parent oysters, and improve shell color and growth traits through population selection to obtain diploid Fujian oyster strains with golden and black shells. b. By combining the additive genetic variance accumulated within the Fujian oyster's golden shell and black shell varieties during the breeding process with the non-additive genetic variance between varieties through inter-strain hybridization, a new diploid Fujian oyster hybrid variety with fast growth rate and high survival rate can be obtained. c. Using a diploid Fujian oyster black-shelled strain as the male and female parents, a triploid Fujian oyster black-shelled strain is induced. Using the triploid Fujian oyster black-shelled strain as the female parent and the diploid Fujian oyster black-shelled strain as the male parent, a tetraploid Fujian oyster black-shelled strain is induced. For the tetraploid Fujian oyster black-shelled strain, individuals with uniform development, ≥15 million eggs, and deep yolk color are selected from the triploid Fujian oyster black-shelled strain as the female parent. Individuals from the diploid Fujian oyster black-shelled strain are selected as the female parent. In the oyster shell black strain, individuals with vigorous sperm motility are selected as the male parent. Fertilization is carried out at a sperm:egg ratio of 50:

1. 8-10 minutes after fertilization, 1 mg / L of cytochalasin B is added to inhibit the release of the first polar body of the fertilized egg. The group without reagent is used as the control group. When 50% of the fertilized eggs in the control group show the first polar body, the eggs are washed. After hatching and selection, the ploidy of the larvae is determined. The group with a tetraploid rate of more than 80% can obtain the tetraploid Fujian oyster shell black induction strain. d. Through population selection and family selection, the shell color, growth and reproductive capacity of the tetraploid Fujian oyster black-shelled strain are selected in a targeted manner to obtain an improved tetraploid Fujian oyster black-shelled strain with pure shell color, fast growth rate and high reproductive capacity. e. Using the new diploid Fujian oyster hybrid strain described in step (b) as the female parent and the tetraploid Fujian oyster shell black improved strain described in step (d) as the male parent, a fast-growing and high-survival-rate triploid Fujian oyster improved strain can be obtained through hybridization. Individuals with uniform development, ≥15 million eggs, and dark yolk color are selected from the new diploid Fujian oyster hybrid strain as the female parent, and individuals with pure black shell color and vigorous sperm motility are selected from the tetraploid Fujian oyster shell black improved strain as the male parent. Fertilization is carried out at a sperm:egg ratio of 50:

1. Larval and adult oyster rearing are carried out using conventional methods to obtain a fast-growing and high-survival-rate triploid Fujian oyster improved strain.

2. The method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster according to claim 1, characterized in that: In step (a), oysters with good vitality, beautiful shell shape and no damage, and both shells are pure gold or pure black are selected as the parent oysters of the two basic populations. When selecting the populations, the top 10% of individuals in terms of shell height and wet weight are selected and continuously selected for 3 to 4 generations of high-intensity breeding.

3. The method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster according to claim 1, characterized in that: In step (b), when hybridization is carried out, individuals with high shell height and total weight are preferred. At least 50 male and female parents are selected from the black-shelled and gold-shelled strains respectively, and fertilization is performed at a sperm:egg ratio of 50:

1.

4. The method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster according to claim 1, characterized in that: In step (c), the triploid Fujian oyster shell black strain is induced by selecting individuals with pure black color, high shell height, and total weight from the diploid Fujian oyster shell black strain as parents. Among them, individuals with uniform development, ≥15 million eggs, and dark yolk color are selected as mothers, and individuals with full gonads and vigorous sperm motility are selected as fathers. Sperm and eggs are obtained by "dry peeling method".

5. The method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster according to claim 1, characterized in that: In step (c), the triploid Fujian oyster shell black strain is induced. After fertilization for 15 minutes, 0.75 mg / L of cytochalasin B is added to inhibit the release of the second polar body. After continuous treatment for 15 minutes, the fertilized eggs are soaked in seawater with 2 mg / L dimethyl sulfoxide solution. After hatching and selection, the ploidy of the larvae is determined. The group with a triploidity rate of more than 95% is retained. The larvae are cultured and the adult oysters are raised using conventional methods to obtain the triploid Fujian oyster shell black induced strain.

6. The method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster according to claim 1, characterized in that: In step (d), when conducting group selection, select black-shelled tetraploid Fujian oysters with pure black shell color, shell height and total weight, and high reproductive capacity as parents. Select the top 10% of individuals in terms of shell height and total weight, and select more than 50 males and 50 females in each generation for 2-3 consecutive generations of high-intensity selection.

7. The method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster according to claim 1, characterized in that: In step (d), when conducting population selection, step (c) is repeated to obtain black-shelled tetraploid Fujian oysters with different genetic backgrounds, which are continuously added to the original tetraploid population to ensure the high genetic diversity of the tetraploid population.

8. The method for cultivating a fast-growing, high-survival-rate triploid improved strain of Fujian oyster according to claim 1, characterized in that: In step (d), when selecting pedigrees, the best parents are tetraploid Fujian oysters with pure black shell color, high shell height and total weight. 40-50 full-sib families are established through one-to-one fertilization. After the gonads of the previous generation of families mature, the families with pure black shell color and a tetraploidity rate of more than 95% are selected as the parents of the next generation of families. Tetraploid individuals from different families are randomly fertilized one-to-one to obtain the next generation of families. After 2-3 generations of pedigree selection, a tetraploid Fujian oyster improved black shell strain with high tetraploidity rate, pure shell color and high reproductive capacity can be obtained.

Images