藏马微卫星标记遗传多样性研究

Development and research of the microsatellite markers of Tibetan horse

为研究西藏藏马种群的微卫星多态性,应用生物素标记的微卫星探针与藏马基因组酶切片段杂交,通过链霉亲和素包被的磁珠捕获150~1 000 bp含有微卫星序列的DNA片段,连接到pEASY-T1载体中,构建藏马基因组微卫星富集文库。结果表明:从2 380个转化子中随机挑取112个阳性克隆进行测序后,经筛选比对发现,从62个微卫星中筛选出39个在马属基因组卫星库中未被检测到的微卫星;成功设计了10对藏马微卫星引物,经聚合酶链式反应-单链构象多态性(PCR-SSCP)方法检测到3个具有多态性的微卫星标记,其中有2个多态性信息含量(PIC)值均大于0.5。因此,这3个微卫星标记可应用于藏马遗传多态性检测、种群遗传结构等方面研究,为藏马遗传连锁图谱的构建、辅助育种、群体遗传学分析、基因组结构分析以及分子进化研究等提供依据。

The aim of this study was to find microsatellite markers of Tibetan horse population in high altitude area of Tibet.The biotin-labeled microsatellite probes were used to hybridize with Tibetan horse genomic fragments.DNA fragments 150-1 000 bp containing microsatellite sequences were captured by streptavidin-coated magnetic beads and ligated into pEASY-T1 vector.An microsatellite sequences enriched insert library was constructed.The results showed that:A total of 112 positive clones were picked from 2 380 transformants for sequencing and 39 microsatellites were indentified from the Equine genomic satellite library out of 62 microsatellites.Ten pairs of Tibetan horse microsatellite primers were successfully designed.Three polymorphic microsatellite markers were detected by PCR-SSCP method,and two of which had PIC values greater than 0.5.In conclusion,the three microsatellite markers could be applied to the study of genetic polymorphism detection of Tibetan horses and the genetic structure of the population.This study provided genetic bases for the linkage map construction,breeding,population genetic analysis,genomic structure analysis and molecular evolution research of Tibetan horse.