合浦珠母贝微卫星DNA标记分离与分析

采用生物素标记的（CA）15、（AG）12、（ACA）15、（GATA）8、（GATT）75个探针和磁珠富集法构建马氏珠母贝（Pinctada martensii Dunker）基因组微卫星富集文库。随机挑选500个进行PCR筛选，得到138（27．6％）个候选克隆，测序分析发现130个克隆含有微卫星基重复单元。进一步通过序列比对，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。获得的微卫星序列中，属于完全型序列的有154条，不完全型重复型序列有19条，复合型重复序列有6条。序列长度为70～490bp，平均295bp。微卫星核心序列两碱基重复3到39次，绝大多数序列重复次数大于10。基于微卫星两端的侧翼序列设计并获得了13对能够在马氏珠母贝基因组有效扩增的微卫星引物。本研究旨为进一步开展马氏珠母贝分子育种及资源评价分析提供基础资料。

Isolation and Analyzing Pinctata fucata with Microsatellite DNA Marker

Microsatellite - enhanced genomic Library of the Pinctada martensii Dunker was constructed using repeat - enrichment method with biotin-labeled oligos （CA）15, （AG）12, （ACA）15, （GATA）8, （GATT） 7] and streptavidin magnetic beads. From a total of randomly selected 500 clones, 138 clones （27.6%） were acquired after PCR selection. Of which, 130 clones are discovered to contain microsatellites. By alignment, 109 special microsatellite clones are confirmed finally. In addition, the microsatenites can be divided into three types of 154 pure sequences, 19 interrupted sequences and 6 compound se- quence. The sequence sizes range from 70bp to 490bp with an average of 295 bp. The repeats of two nucleotides within core se- quences are 3 to 39 times, and most of them are more than 10 times. Thirteen pairs of primers were designed and effective for PCR amplification. This study provides a base for molecular breeding and assessment of germplasm resources of the Pinctada martensii Dunker.