| In the late 20th, Haliotis diversicolor become the most important farming abalone in southern China because of its rapid growth. As the marine environment, a single breeding population genetic structure, increase the probability of inbreeding, lack of fast growth, good quality and strong resistance new abalone breeding. Meanwhile, over-farmed, farming pollution, frequent red tides external factors is a serious constraint abalone aquaculture sustainable development. How to enhance the resilience of abalone and improve its quality and quality improvement of germplasm resources situation has become an important research topic in China. With molecular biology techniques developed molecular markers not only for the investigation of genetic diversity and population genetic structure, but also for germplasm evaluation, identification, traceability provides the conditions,also lay the foundation for Marker-assisted breeding system. In the present study, a panel of microsatellite markers were isolated and characterized, and analysis of cultured abalone populations genetic diversity. The results obtained here are as follows:(1) As the Haliotis diversicolor in the public database of gene sequence information in relatively small, particularly EST sequences, and the Haliotis diversicolor of microsatellite markers research was virtually nonexistent. A complementary DNA (cDNA) library with total 8016 expressed sequence tags (ESTs) was constructed and sequenced from whole body of Haliotis diversicolor Reeve. A total of 8016 expressed sequence tags (ESTs) and clustered into 1087 contigs and 3694 singletons. Among the 4781 unique genes identified, 1882 (23.5%) genes had no significant matches to known sequences in public database. The remaining 2899 (36.2%) genes that exhibited homology with gnese of known functions. 161 ESTs sequences (3.72%)contain 173 microsatellite were found.(2) 4781 unigenes were assembled from this dataset and screened for simple repeat sequences (SSRs) with at least 5 repeats for di-, tri-, tetra- and penta-, and 4 repeats for hexanucleotide repeats. Total 173 SSRs were found, and the number of di- and trinucleotide repeats are 110 (63.6%) and 47 (27.2 %) separately. Primers were designed according 93 SSR-containing ESTs with sufficient flanking sequences to acquire 78 (83.9%) successful polymerase chain reactions (PCRs) including 25 (32.1%) products longer than expected sizes probably due to introns. 53 pairs that produced expected fragments were screened for polymorphism between eight abalone individuals from three populations. 19 (35.8%) of them with polymorphism were examined in one population (40 individuals) to test the number of alleles. The number of alleles ranged from 2 to 16 with an average of 7.84. The observed heterozygosity ranged from 0.412 to 0.748 , and the expected heterozygosity ranged from 0.653 to 0.846.(3)Genetic diversity of 7 cultured populations of the Haliotis diversicolor from Shantou, Shanwei, Huidong, Shenzhen, Zhanjiang, Xuwen (above all of Guangdong Province, China) was analyzed using seven microsatellite markers (ZSB2,ZSB4,ZSB6,ZSB8,ZSB10,ZSB13,ZSB17). Seventy-six alleles of these seven microsatellite loci were found in these seven populations. The observed number of alleles of these microsatellite loci was 8-16 in each population with the average number of alleles was 10.87, and the average effective number of alleles was 3.8-10.8. The observed population heterozygosity was 0.472-0.914 and the observed expected heterozygosity was 0.748-0.920. The polymorphism of the seven populations were all high, and the PIC of the pupations were 0.645-0.889 ( PIC >0.5). Compared two populations of wild domesticated breeding, the five breeding groups were of different degrees of overall reduction in genetic diversity. In this study, Fst values range from 0.013-0.167. Between the two wild domesticated breeding populations and between wild domesticated breeding and cultured populations Fst values are greater than 0.08, indicating that between wild domesticated breeding populations, and between wild domesticated breeding populations and cultured populations exist significant genetic differentiation. The results did not show the consistent relationship between the geographic and the genetic distances, suggesting the existence of exchanges of breeds and eggs between the hatcheriesthe abalone.Generally speaking, groups of abalone farming is still a high level of genetic diversity, these results provided a useful information for the selective breeding, germplast conservation and its application of Haliotis diversicolor. |
