Construction And Application Of Genetic Linkage Map For Pacific Abalone (Haliotis Discus Hannai) Based On Microsatellite And SNP Markers

A total of 301 novel microsatellite markers (SSR) of the Pacific abalone were developed by microsatellite-enriched libraries and Expressed Sequence Tag (EST) database mining. The polymorphisms of 245 markers were characterized by genotyping a wild population comprising 37 unrelated individuals. The result showed the number of allele ranged from two to 18, and the values of expected and observed heterozygosities ranged from 0.053 to 0.873 and from 0.054 to 1.000, respectively. EST database mining was proven to be efficient and low-costly approach to obtaining new microsatellite markers, and microsatellite enrichment libraries construction was efficient and suitable to develop a large number of microsatellite markers.Fifteen single nucleotide polymorphism (SNP) markers were developed by EST database mining and PCR direct sequencing. The PCR Primers were designed by the method of polymerase chain reaction amplification of multiple specific alleles (PMASA). The polymorphisms of 15 markers were assessed with 37 unrelated individuals and the result showed that the minor allele frequency ranged from 0.1216 to 0.4865. For the fourteen nuclear SNPs, the value of observed heterozygosity ranged from 0.189 to 0.865, while the expected heterozygosity ranged from 0.217 to 0.507. Only one locus (HdS82) deviated significantly from Hardy-Weinberg equilibrium after Bonferroni correction.These polymorphic markers presented in this study provide a useful tool for population genetics, pedigree analysis, linkage map construction and marker-assisted selection (MAS) of H. discus hannai.The genetic linkage map of H. discus hannai was constructed using 342 microsatellite markers, 11 SNP markers and one shell color marker. Linkage mapping was performed using two F1 outbred families, and the integrated linkage map was generated by incorporating map information from the two families. The integrated linkage map contained 324 markers covering 810.32 cM with an average spacing of 2.65 cM and 88.8% of genome coverage. The number of linkage groups in the integrated linkage map was 18, which was consistent with the haploid chromosome number of H. discus hannai. The shell color marker was located in LG 9. The detection and location of QTLs were performed based on the composite linkage map. Ten putative QTLs (LOD≥3.8) associated with the growth of Pacific abalone were detected and located in the composite linkage map, among which three QTLs for the shell length, two for the shell width, two for the gross weight and four for the flesh weight, respectively. They were scattered on LG5 and LG6, explaining 13.4% to 29.1% of the trait variation. A total of 14 functional genes were located in the composite linkage map through the molecular markers. The location of functional genes and QTLs associated with the growth would be very useful for molecular marker-assisted selection and map-based cloning of functional genes.