采用高通量技术开发花鲈二碱基重复微卫星标记

Development of Dinucleotide Microsatellite Loci for Lateolabrax maculates Using Next Generation Sequencing Technology

花鲈是中国重要的捕捞和养殖对象之一。本研究利用高通量测序法开发花鲈微卫星标记,共获得60632个二至六碱基重复微卫星序列,从52747个二碱基重复序列中随机挑选150个位点进行引物合成及多态性检测,最终开发出27个具有多态性的微卫星标记。群体遗传学检测结果显示,27个微卫星标记的等位基因数为6~22(均值12.667),观测杂合度(Ho)为0.323~1.000(均值0.614),期望杂合度(He)为0.723~0.934(均值0.861),多态信息含量(PIC)为0.665~0.915(均值0.830),各位点PIC值均大于0.500,表明所开发的二碱基重复微卫星标记均具有较高的多态性。哈迪-温伯格平衡(HWE)检测结果显示,14个标记符合HWE,其中LM2-11与LM2-2、LM2-16之间存在连锁不平衡(p<0.050),其他符合HWE的标记间不存在连锁不平衡。Wilcoxon检验结果显示,花鲈群体并未偏离L-shaped分布,表明其经历过近期的遗传瓶颈效应。本研究开发的27个微卫星位点可为花鲈的分子标记辅助育种、增殖放流遗传效应评估、群体遗传学等研究提供有效的分子标记。

The spotted sea bass(Lateolabrax maculates)is an important species for fishing and culture in China.In this study,a total of 60632 microsatellites(di-to hexanucleotide repeats)were developed by next generation sequencing technology.One hundred and fifty microsatellite loci were randomly selected for amplification and polymorphism analysis from 52747(87%)dinucleotide repeat,of which 27 were polymorphic and well-resolved.The population genetic analysis showed a high polymorphism with a range of 6 to 22 alleles(mean 12.667).Observed and expected heterozygosities varied from 0.323 to 1.000(mean 0.614)and 0.723 to 0.934(mean 0.861),respectively.The polymorphism information content(PIC)ranged from 0.665 to 0.915(mean 0.830),and the PIC at each loci is greater than 0.500,indicating that the developed dinucleotide microsatellite markers were highly polymorphic.The Hardy-Weinberg equilibrium test showed that 14 microsatellite markers conformed to HWE.Among 14 loci,a linkage imbalance existed between LM2-11 and LM2-2,LM2-11 and LM2-16(p<0.050),and there is no linkage imbalance between other markers.Wilcoxon test revealed that the population did not deviate from the L-shaped distribution,indicating that it had experienced the recent genetic bottleneck effect.The present microsatellites would be useful for further molecular marker-assisted breeding,genetic effect assessment of stock enhancement,and population genetic studies for L.maculates.