Hybrid breeding method of southern and northern populations of Ruditapes philippinarum

Abstract

The invention relates to a hybrid breeding method for a southern group and a northern group of Manila clams, and belongs to the technical field of shellfish genetic breeding in aquaculture. The method comprises the following steps: collecting a northern group and a southern group, taking shell length as a breeding index, selecting the top 10% of individuals in shell length as parents, carrying out fast growth breeding in the southern group and the northern group respectively, obtaining the breeding F1 generation of the southern group and the northern group respectively, and cultivating them to sexual maturity; obtaining the breeding F2 generation of the southern group and the northern group respectively by the same breeding method, selecting well-developed individuals as candidate parents, placing them in the same indoor cultivation pool for synchronous maturation; artificially inducing spawning of parent shellfish, and adopting methods including freshwater flushing, high-concentration algae water feeding and air flow stimulation; and obtaining hybrid offspring showing obvious hybrid advantages in growth and survival.

Description

Technical Field

[0001] The invention belongs to the technical field of shellfish genetic breeding in aquaculture, and specifically relates to a hybrid breeding method of a southern group and a northern group of Manila clams, so as to cultivate a new clam variety (line) suitable for breeding in both southern and northern my country, with fast growth rate and high survival rate. Background Art

[0002] Hybrid breeding is a widely used method in aquatic animal breeding. It can obtain hybrid offspring with different gene combinations by hybridizing different parents. Especially in shellfish breeding, intraspecific hybridization and interspecific hybridization are considered to be one of the effective means to cultivate excellent varieties. When conducting hybrid breeding, the selection of parents is very critical. Not any hybridization of two populations can obtain hybrid vigor. Therefore, it is particularly important to examine the heritability of the traits of the parents. Parents should have certain genetic differences and should show high complementarity and heterozygous advantages. By examining and evaluating the traits of the parents (including body shape, growth rate and adaptability, etc.), kinship and geographical distribution, parents suitable for hybridization can be better selected.

[0003] When breeding shellfish, hybridization between different populations can produce more genetic variation, thereby improving the adaptability and productivity of offspring. In addition, during the process of shellfish hybridization, the hybrid offspring eliminates bad genes and improves the expression of good genes through the rearrangement of gene combinations, obtains hybrid vigor, and thus cultivates new shellfish varieties with excellent traits.

[0004] The Manila clam (Ruditapes philippenarum) is one of the four major cultured shellfish in my country. It is widely distributed in the coastal areas of the north and south of my country. It grows fast, has strong adaptability, and has a short culture cycle. It is an excellent shellfish suitable for high-density culture. However, in recent years, with the rapid expansion of Manila clam culture, problems such as slow growth, low meat yield, and high mortality in summer have begun to appear, which seriously restricts the healthy development of its culture industry. In the process of Manila clam culture, slow growth and summer death often occur. Therefore, cultivating new varieties with fast growth and strong resistance is the key to achieving healthy and sustainable development of the clam culture industry. Summary of the invention

[0005] The invention aims to provide a hybrid breeding method of a southern group and a northern group of Ruditapes philippinarum.

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

[0007] A hybrid breeding method of a southern population and a northern population of Ruditapes philippinarum, the method being specifically as follows:

[0008] Step 1: Collect northern and southern populations, use shell length as a breeding index, select the top 10% of individuals in shell length as parents, carry out fast growth breeding in the southern and northern populations, obtain breeding F1 generations of the southern and northern populations, and cultivate them to sexual maturity;

[0009] Step 2: Using the same breeding method as step 1, obtain the F2 generation of the southern group and the northern group respectively, select well-developed individuals as candidate parents, and place them in the same indoor cultivation pool for synchronous ripening;

[0010] Step 3: Artificial induction of spawning of broodstock, using methods including freshwater flushing, feeding with high-concentration algae water and air flow stimulation;

[0011] The fresh water flushing is to take out the broodstock to be produced from the culture pond and use fresh water to fully flush the broodstock for 10-15 minutes;

[0012] The high-concentration algae water feeding is to slowly pour the high-concentration Chaetoceros algae water around the parent shellfish to be produced to promote the parent shellfish's feeding, and the high-concentration Chaetoceros algae is 2-4 million / ml;

[0013] The airflow induction of labor is to open the oxygenation tube in the induction pool and place the air stone near the parent shellfish to speed up the surrounding water flow;

[0014] Step 4: The fertilized eggs are transferred to an incubation bucket for continuous aeration incubation. After hatching, the larvae are fed with golden algae and horn algae in a gradual manner.

[0015] Furthermore, in the first three days after the larvae were hatched, golden algae were fed according to the standard of 4000-6000 algae / ml, and then the proportion of Chaetoceros algae began to be gradually increased, and the feeding ratio was 1:4 for Chaetoceros algae: golden algae, gradually increased to 1:3, 1:2, and 1:1. After one week, all feeding was changed to Chaetoceros algae; the larval breeding density was maintained at 8-10 / mL, the water temperature was 18-25°C, the pH was 7.8-8.2, and the salinity was 19-27psu.

[0016] Compared with the prior art, the present invention has the following advantages:

[0017] First, the southern clam population collected by the present invention has significant genetic differentiation and a large genetic distance from the northern clam population. By carrying out two consecutive generations of growth and breeding within the southern population and the northern population, the F2 generation of the southern population and the northern population with excellent traits is obtained, and the hybrid offspring of the southern population and the northern population show obvious hybrid advantages in growth and survival.

[0018] Secondly, the large temperature difference between the north and south of my country has resulted in the northern and southern groups of clams having their own advantages in terms of environmental adaptability. The northern group is more adaptable to low-temperature environments, while the southern group grows in warmer waters and is more adaptable to high-temperature environments. Therefore, the hybrid offspring of the southern and northern groups have a wider temperature range and higher environmental adaptability.

[0019] In addition, clams are dioecious shellfish, and the parents' spermatozoa and spawning are not synchronous. The present invention places the southern breeding population and the northern breeding population in the same indoor cement pool for synchronous ripening and induced spawning, picks out the parents who ejaculate or lay eggs, and uses fresh water flushing, high-concentration algae water feeding and air flow stimulation to stimulate the male and female parents to spawn and ejaculate synchronously. At the same time, fresh water is used to flush the shell surface and sperm and eggs are collected separately, which effectively avoids cross contamination. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 This is a statistical chart of the survival rate of the early stages of clam self-pollination and hybrid offspring. DETAILED DESCRIPTION

[0021] The technical solution of the present invention is further explained below by way of embodiments, but the protection scope of the present invention is not limited in any form by the embodiments.

[0022] Example 1

[0023] A hybrid breeding method of a southern population and a northern population of Ruditapes philippinarum, the method comprising the following specific steps:

[0024] 1) Clam populations from Laizhou Bay, Shandong (northern population) and Zhanjiang, Guangdong (southern population) were collected, and shell length was used as the breeding index. The top 10% of individuals in shell length were selected as parents, and fast growth breeding was carried out within the southern and northern populations to obtain the F1 generation of the southern and northern populations, respectively. When the juveniles had a shell length of 8-10 mm, they were cultured on the mudflats until sexual maturity.

[0025] 2) Using the same breeding method, select the F1 generation with mature gonads, select the individuals in the top 10% of shell length as parents, and obtain the F2 generation of the southern group and the northern group. After bred to sexual maturity, select the individuals with large size and full gonads as candidate parents. The magnetic parents and male parents of the southern group and the northern group are placed separately in a birthing basket. A total of 6 birthing baskets are placed in the same cement pool for indoor synchronous ripening, and the water temperature is controlled below 17°C.

[0026] 3) Artificially induce spawning of the parents by means of freshwater flushing, feeding with high-concentration algae water, and air flow stimulation;

[0027] The freshwater flushing is to take out the broodstock to be born in the induced spawning pool and place them on flat ground, then open the freshwater valve to fully flush and moisturize the broodstock for 10-15 minutes, which can stimulate the spawning and spermatogenesis of the broodstock.

[0028] The high-concentration algae water feeding is to prepare a 20-liter bucket to take high-concentration (about 2 million / ml) Chaetoceros algae water and slowly pour it around the parent shellfish to be produced. This will promote the parent shellfish's feeding and stimulate the parent shellfish to lay eggs and ejaculate.

[0029] The air flow stimulation is to open the oxygenation tube in the induced labor pool, set the oxygenation tube near the parent shellfish to be born, and then put the air stone of the oxygenation tube into the birthing basket (the specifications of the perforated plastic basket are 1.2m long×0.6m wide×0.2m high), and place 2 air stones in each birthing basket. The specifications of the air stones are 3cm in diameter and 4cm in height, so as to speed up the surrounding water flow and continuously stimulate the parent shellfish to lay eggs and ejaculate.

[0030] During the night induction of spawning, the experiment of inducing spawning and ejaculation of the broodstock in the spawning frame was carried out. The broodstock were divided into three groups and different methods or combinations of methods were used to induce spawning and ejaculation. The details are as follows:

[0031] The first group of parent pigs used only air flow stimulation and began to lay eggs and ejaculate in about 5 hours, accounting for about 65% of the output.

[0032] The second group of broodstock were fed with high-concentration algae water and stimulated by airflow. They began to spawn and ejaculate between 2.5 and 3 hours, accounting for about 70% of the output.

[0033] The third group of broodstock used a combination of freshwater flushing, high-concentration algae water feeding, and air flow stimulation. They successfully laid eggs and ejaculated in about an hour, with an output ratio of about 75%.

[0034] When some individuals started to lay eggs and ejaculate, male and female individuals were selected from the southern breeding parents and the northern breeding parents respectively. The shell surfaces were rinsed with fresh water to prevent cross contamination and placed in separate sperm and egg collection buckets to obtain sperm and eggs from the southern breeding group (Z♂ and Z♀) and sperm and eggs from the northern breeding group (L♂ and L♀).

[0035] 4) The four groups of sperm and eggs (L♀, L♂, Z♀ and Z♂) collected were artificially inseminated by the double-row hybridization method to establish self-pollinated groups (northern inbred line LL: L♀×L♂; southern inbred line ZZ: Z♀×Z♂) and hybrid groups (orthogonal LZ: L♀×Z♂ and reciprocal ZL: Z♀×L♂) of the northern and southern breeding populations, and to obtain self-pollinated and hybrid progeny of the northern and southern breeding populations.

[0036] 5) The fertilized eggs were transferred to a 50L large white barrel for continuous aeration incubation. After the hatched larvae were selected, 3 replicates were set up in each group. At different developmental stages, the shell length growth and survival rate were measured regularly. The larval cultivation density was maintained at 8-10 / mL, the water temperature was 18-25℃, the pH was 7.8-8.2, and the salinity was 19-27psu. During the cultivation of each group of larvae, the water was completely changed every other day. In the first three days after the larvae were hatched and selected, golden algae were fed according to the standard of 4000-6000 algae / ml, and then the proportion of horn algae was gradually increased. The feeding ratio was horn algae: golden algae was 1:4, and gradually increased to 1:3, 1:2, and 1:1. After one week, all were changed to horn algae; feeding was carried out daily according to the actual feeding situation of the larvae.

[0037] 6) By measuring the shell length of larvae and juveniles in the self-pollination group and the hybrid group, it was found that the shell length of the orthogonal group LZ and the reciprocal group ZL was significantly higher than that of the two self-pollination groups after the 15th day. The growth rate of the orthogonal group LZ was 28.36% higher than that of the self-pollination group LL, and the growth rate of the reciprocal group ZL was 34.33% higher than that of the self-pollination group LL. Among them, the heterosis rate of the shell length of the reciprocal group ZL was the highest on the 20th day, which was 33.42%. By comparing the survival rates of each group on the 5th, 10th, 15th, and 20th days, it was found that the survival rate of the hybrid group was significantly higher than that of the self-pollination group ( Figure 1 On the 20th day, the survival rate of the reciprocal cross group ZL was the highest (40%), followed by the survival rate of the orthogonal cross group LZ (28%), but still much higher than the survival rates of the self-cross groups LL and ZZ (6.67% and 11.00%).

Claims

1. A hybrid breeding method of southern and northern populations of Ruditapes philippinarum, characterized in that: The method is specifically as follows: Step 1: Collect northern and southern populations, use shell length as a breeding index, select the top 10% of individuals in shell length as parents, carry out fast growth breeding in the southern and northern populations, obtain the breeding F1 generation of the southern and northern populations, and cultivate them to sexual maturity; Step 2: Using the same breeding method as step 1, obtain the F2 generation of the southern group and the northern group respectively, select well-developed individuals as candidate parents, and place them in the same indoor cultivation pool for synchronous ripening; Step 3: Artificial induction of spawning of broodstock, using methods including freshwater flushing, feeding with high-concentration algae water and air flow stimulation; The fresh water flushing is to take out the broodstock to be produced from the culture pond and use fresh water to fully flush the broodstock for 10-15 minutes; The high-concentration algae water feeding is to slowly pour the high-concentration Chaetoceros algae water around the parent shellfish to be produced to promote the parent shellfish's feeding, and the high-concentration Chaetoceros algae is 2-4 million / ml; The air flow stimulation is to open the oxygenation tube in the oxytocin pool and place the air stone near the parent shellfish to speed up the surrounding water flow. Step 4: The fertilized eggs are transferred to an incubation bucket for continuous aeration incubation. After hatching, the larvae are fed with golden algae and horn algae in a gradual manner.

2. The method according to claim 1, characterized in that In the first three days after the larvae hatched, they were fed with golden algae at a standard of 4000-6000 algae / ml. Then the proportion of Chaetoceros algae was gradually increased, and the feeding ratio was 1:4 for Chaetoceros algae: golden algae, and gradually increased to 1:3, 1:2, and 1:

1. After one week, all feeding was changed to Chaetoceros algae. The larval breeding density was maintained at 8-10 / mL, the water temperature was 18-25℃, the pH was 7.8-8.2, and the salinity was 19-27 psu.

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