合浦珠母贝微卫星DNA标记的分离与筛选

目的]研究合浦珠母贝微卫星DNA标记的分离与筛选。方法]采用生物素标记的5个探针，用磁珠富集法构建合浦珠母贝基因组微卫星富集文库，并对文库进行筛选、测序与引物多态性筛选。结果]对500个克隆进行PCR筛选，测序分析发现130个克隆含有微卫星重复单元。序列比对后最终获得109个具有特异微卫星序列的阳性克隆，其中包含179个微卫星DNA结构域。基于微卫星两端的侧翼序列设计并获得了13对能够在合浦珠母贝基因组有效扩增的微卫星引物，其中8对在种群中（n=32）具有扩增多态性，其多态信息含量PIC值在0．239～0．787；等位基因数在2～9个；观测杂合度介于0．22～0．56；期望杂合度的变化范围为0．25～0．83。结论]这研究为进一步开展合浦珠母贝的遗传分析提供了基础资料。

Isolation and Screening of Microsatellite DNA Markers from Pinctada fucata

Objective] The purpose was to study the isolation and screening of microsatellite DNA markers from Pinctada fucata.Method] Microsatellite-enhanced genomic library of the P.fucata was constructed by using magnetic bead-enrichment method with 5 biotin-labeled probes and made for screening,sequencing and primer polymorphic screening.Result] From a total of randomly selected 500 clones,130 clones were discovered to contain microsatellites through sequencing analysis.By sequence alignment,109 special microsatellite positive clones were confirmed finally,which contained 179 microsatellite DNA domains.Based the side sequence design of microsatellite,13 pairs of primers that were effective for PCR amplification in P.fucata genome were obtained,in which,8 loci showed the polymorphism in a population(n=32),with their PIC values from 0.239 to 0.787,the allele number from 2 to 9,the observed heterozygosity from 0.22 to 0.56 and the expected heterozygosity from 0.25 to 0.83.Conclusion] This study provided the basic data for further studying the genetic analysis of P.fucata.