许氏平鲉(Sebastes schlegelii)EST-SSR标记开发及通用性检测

利用已发表的EST序列设计引物并结合PCR筛选的方法,对许氏平鲉多态EST-SSR引物进行了开发。结果表明:(1)许氏平鲉EST-SSR位点发现率为9.14%,核心序列以二碱基重复出现频率最高,占到33.03%,其中以TG/CA基序最为丰富。(2)共设计引物56对,46对实现有效扩增,其中18对具有多态性。(3)野生群体遗传多样性分析显示,每个位点观测等位基因数(Na)为2—9,平均观测等位基因数为3.89;观测杂合度(Ho)、期望杂合度(He)及平均值分别是0.0333—0.8000、0.0333—0.7927和0.3037、0.3757;每个位点的多态信息含量(PIC)在0.0323—0.7522之间,其中高度多态位点5个,中度多态位点5个;经Bonferroni校正后,3个位点显著偏离Hardy-Weinberg平衡,但两两位点间不存在连锁不平衡现象。(4)许氏平鲉多态EST-SSR引物对朝鲜平鲉、褐菖鲉通用性检测结果表明,引物对二者的通用率和多态率分别为88.89%、100.00%和33.33%、88.89%。研究表明,这些通用引物可用于相关物种的遗传多样性评价、系统进化分析和比较基因组作图等研究。

DEVELOPMENT OF EST-SSR MARKERS FOR ROCKFISH SEBASTES SCHLEGELII AND CROSS-SPECIES AMPLIFICATION

We designed primers with published expressed sequence tag(EST) sequences in combination with PCR amplification, polymorphic EST-SSR(simple sequence repeats) markers in rockfish Sebastes schlegelii. The results show that the discovery rate of EST-SSR on EST sequences was 9.14%; dinucleotide motif had the highest frequency at 33.03%; and TG/CA motif was the richest one. In total, primers, 46 pairs out of 56 pairs that designed proved effective, and 18 pairs expressed polymorphism. We then applied these loci to analyze 30S. schlegelii DNA specimens in population diversity. The observed number of alleles(N_a) ranged 2-9 in average of 3.89. The observed heterozygosity(H_o), expected heterozygosity(H_e) and their average values were 0.0333-0.8000, 0.0333-0.7927, 0.3037 and 0.3757, respectively. Polymorphic information content(PIC) ranged 0.0323-0.7522, five loci were highly polymorphic, and 4 were moderately polymorphic. After Bonferroni correction, 3 loci deviated significantly from Hardy-Weinberg Equilibrium(P-value), but no linkage disequilibrium phenomenon. The 18 loci were surveyed in S. koreanus and Sebastiscus marmoratus, the cross-species proportions and polymorphic rates were 88.89%, 100.00% and 33.33%, 88.89%, respectively. Therefore, these EST-SSR loci of S. schlegelii may facilitate studies on genetic diversity, phylogeny, and genomes mapping of related species.