日本沼虾多态性标记筛选及群体遗传结构分析

Screening of Polymorphic Markers and Analysis of Population Genetic Structure of Oriental River Prawn Macrobrachium nipponense

从转录组测序数据中筛选重复次数较多、两翼序列较长的简单重复序列用于多态性EST-SSR标记的筛选,利用荧光标记SSR技术分析太湖、长江下游(崇明岛附近)、微山湖和东平湖4个地理群体日本沼虾的遗传多样性。从日本沼虾转录组测序数据中选择546个cDNA序列,设计了60对引物,有33对能扩增出稳定、清晰的条带,其中12对呈现出良好的多态性。这12个多态性微卫星标记的等位基因数为3~7,观测杂合度为0.1724~0.9630,期望杂合度为0.1942~0.8462。群体遗传多样性分析结果显示,微山湖群体的平均期望杂合度和观测杂合度最高,分别为0.8403、0.8350;东平湖的次之,为0.8350、0.7854;长江下游群体的最低,为0.7145、0.7909。由此可见,北方2个群体的遗传多样性要高于南方2个群体。4个日本沼虾群体的分子方差分析显示,东平湖群体与长江下游群体、微山湖群体与长江下游群体的遗传分化指数介于0.05~0.15,达到中等分化程度。聚类分析显示,微山湖和东平湖群体聚类为一支,再先后与太湖群体、长江下游群体(崇明岛附近)聚类,这与各日本沼虾群体实际的地理距离吻合,反映出日本沼虾的生物地理进化关系。

In the present study,novel EST-linked polymorphic microsatellite markers were isolated and characterized from the transcriptome data in oriental river prawn Macrobrachium nipponense,and the genetic structure was analyzed in four populations of oriental river prawn collected from Weishanhu Lake,downstream of Changjiang River,Taihu Lake,and Dongpinghu Lake using fluorescence SSR.A total of 546 cDNA sequences containing microsatellite were selected and 60 pairs of primers were designed,in which 33 pairs of primers were amplified clearly and stably,with 12 of polymorphic pairs of primers.The number of alleles per locus were ranged from 3 to 7 in the 12 isolated polymorphic loci,with average observed and expected heterozygosity per locus ranged from 0.1724 to 0.9630 and from 0.1942 to 0.8462,respectively.The genotyping of 88 individuals(22 individuals each stock)were found to be the mean expected heterozygosity of 0.8403 and observed heterozygositiy of 0.8350 in Weishanhu Lake population,0.8350 and 0.7854 in Dongpinghu Lake,0.7399 and 0.8134 in Taihu Lake,and 0.7145 and 0.7909 in downstream population of Changjiang River,the higher genetic diversity of population in North China than those in South China.The analysis of molecular variance indicated that the FST value between Dongpinghu Lake population and downstream population of Changjiang River,and between Weishanhu Lake population and downstream population of Changjiang River was higher than 0.05 and lower than 0.15,showing a moderate differentiation level,with genetic distances ranging from 0.0158 to 0.0384.The UPGMA tree showed that populations of Dongpinghu Lake and Weishanhu Lake were clustered into an independent clade,then clustered with Taihu Lake population,and downstream population of Changjiang River in the end,suggesting that their genetic relationship is in accordance with regional distribution.