苏皖地区中华绒螯蟹养殖群体微卫星遗传多样性的评估

Assessment of microsatellite genetic diversity of cultured Eriocheir sinensis populations from Jiangsu and Anhui

中文关键词: 中华绒螯蟹  遗传多样性  微卫星  养殖群体

英文关键词: Eriocheir Sinensis  Genetic diversity  Microsatellite  Cultured population

中文摘要:

      为评估奇数年江苏和安徽中华绒螯蟹(Eriocheir sinensis)养殖群体的遗传背景，进而为后续的遗传改良及新品种培育提供科学依据，本研究利用10对微卫星标记分析了6个中华绒螯蟹养殖群体的遗传多样性和遗传结构。结果显示，所有群体的遗传多样性水平都较高且相近(等位基因数Na=16.0~18.4，有效等位基因数Ne=10.1~12.4，观测杂合度Ho=0.759~0.836，期望杂合度He=0.897~0.916，多态信息含量PIC=0.870~0.892)。群体间遗传距离Dn (0.154~0.277)、遗传分化系数Fst (0.001~0.011)均较小，分子方差分析(AMOVA)结果中群体间遗传变异仅占0.47%，一致表明群体间不存在显著遗传分化。群体间系统发育树显示，6个养殖群体具有共同的祖先型，高淳群体与其他群体亲缘关系最远，结合其高的遗传多样性水平，高淳群体可作为选育基础群之一。Structure遗传结构分析显示，每个养殖群体的遗传组成多样且比例近似，结合近交系数和瓶颈效应分析表明，中华绒螯蟹养殖群体存在较大程度的外源种质混杂。综上，苏皖地区中华绒螯蟹养殖群体遗传多样性仍然较高，具有潜在的开发与利用价值，但其可能存在种质混杂，在开展后续的良种选育时，需对养殖群体进行进一步的研究，提纯种质，使其种质资源得到合理的可持续性利用。

英文摘要:

      Eriocheir sinensis, commonly known as the Chinese mitten crab, is widely distributed in China from the Liaohe River in the north to the Pearl River in the south and in the shallow waters of the estuary into the sea, among which the population in the Yangtze River system has the largest yield and the most delicious taste, and is favored by domestic consumers. It is an important breeding species in the Chinese fishing industry. Since artificial breeding began in the 1980s, the production of Chinese mitten crabs has increased, and this species has become one of the most economically important crabs in the world and the most economically important freshwater crab in China. From the perspective of aquaculture areas, the Yangtze River Basin region accounts for over 80% of the total output in China, with Jiangsu and Anhui provinces ranking first and third, respectively. Therefore, the culture of Chinese mitten crabs in Jiangsu and Anhui plays a pivotal role in the country. After more than 30 years of development, China has established ecological and large-scale breeding systems for juvenile crabs. Therefore, artificial seedlings dominate the market but are mainly distributed in coastal areas such as Jiangsu. However, to reduce production costs, some breeding farms blindly introduce Chinese mitten crabs to one another, and do not perform germplasm detection during breeding, causing germplasm resources to be random and mixed, which directly affects the quality of juvenile crabs and results in the risk of germplasm decline. To evaluate the genetic background of cultured Chinese mitten crab populations in Jiangsu and Anhui in odd-number years, and to provide a scientific basis for subsequent genetic improvement and breeding of new varieties, 10 pairs of highly polymorphic microsatellite markers (simple sequence repeats) were used to analyze the genetic diversity and genetic structure of six cultured populations with 177 individuals in total. Each population contained 28–30 adult individuals with an approximate sex ratio. The leg muscle of the sampled crabs was extracted, and genomic DNA was extracted according to the instructions of the genome extraction kit (TianGen DP304). The microsatellite primers were modified using fluorescent labels (FAM and HEX) prior to PCR, and the PCR products were subjected to capillary electrophoresis. Genemarker 2.2 software was used to interpret typing results. The number of alleles (Na), effective number of alleles (Ne), expected heterozygosity (He), observed heterozygosity (Ho), inbreeding coefficient (Fis), and genetic distance (Dn) were calculated using Popgene 1.32 software, and Hardy-Weinberg equilibrium analysis was performed using the Markov chain method. PIC_CALC 0.6 software calculates polymorphism information content (PIC). Analysis of molecular variance (AMOVA) and the coefficient of genetic differentiation (Fst) were performed using Arlequin 3.5 software. A UPGMA tree was constructed based on inter-population genetic distance using MEGA 4.0 software. According to the frequency of alleles, mutation-drift equilibrium was detected using Bottleneck software. Genetic structure analysis was performed using the Structure 2.3.4 software. The results showed that the genetic diversity levels of the six cultured populations of E. sinensis were high and similar (Na=16.0–18.4, Ne=10.1–12.4, Ho=0.759–0.836, He=0.897–0.916, PIC=0.870–0.892). Genetic distance among populations (0.015–0.277) and the coefficient of genetic differentiation (Fst: 0.001–0.011) were low, and genetic variation among populations accounted for only 0.47% of the total variation in AMOVA. These analyses consistently indicated no significant genetic differentiation among the populations. The phylogenetic tree showed that the six cultured populations had a common ancestor and that the Gaochun population had further relationships with the other populations. Owing to its high genetic diversity, the Gaochun population can be used as a base population for mass selection. Genetic structure analysis showed that the genetic composition of each cultured population was diverse, and the proportions were similar. In this study, the inbreeding coefficient (Fis) of 10 loci in six populations had 43 positive values and 17 negative values, and there were both positive and negative values in each population, indicating that there was not only inbreeding to a certain extent but also a small amount of distant breeding in the six cultured populations of Chinese mitten crabs. The results of genetic structure analysis showed that the number of optimal genetic cluster groups was K=3. However, the chaotic genetic structures among the three groups failed to gather into a relatively independent group. Furthermore, each population had a genetic composition similar to those of the three genetic lineage sources. Considering the high level of genetic diversity in these populations, it is possible that the parental population sources are more complex. This indicated that the cultured population of Chinese mitten crabs had a large number of exogenous genetic hybrids. In conclusion, the genetic diversity of cultured Chinese mitten crab populations in the Jiangsu and Anhui areas is still high, which has potential development and utilization value, but there may be germplasm confounding. Therefore, it is necessary to conduct further studies on the breeding population during the subsequent breeding of improved varieties, such as using molecular markers with high sensitivity that can be used to identify different water systems to analyze the sources of mixed germplasm, and conducting germplasm detection before breeding to purify the germplasm and make reasonable and sustainable use of its germplasm resources.