地黄微卫星富集文库构建及特性分析  

Construction and Characterization of Enriched Microsatellite Library from Rehmannia glutinosa

作　　者：程月琴[1] 焦振彬[1] 张佩[1] 叶永忠[2] 王红卫[1] 

机构地区：[1]河南农业大学植物保护学院,河南郑州450002 [2]河南农业大学生命科学院,河南郑州450002

出　　处：《种子》2013年第5期12-16,共5页Seed

基　　金：国家自然科学基金项目(编号:31170351);河南省教育厅自然科学基金项目(编号:2008A180012)

摘　　要：应用Dynal磁珠-生物素标记的微卫星探针(AC)8,(AG)8和(ATG)12与地黄基因组DNA酶切片段杂交,捕获含有微卫星序列的DNA片段,连接到pMD 18-T载体上,转入感受态细胞Trans 5α,构建地黄富集微卫星文库。利用M 13 F和M 13 R载体序列引物筛选文库,对插入片段长度为400～800 bp的克隆进行测序。共获得96条序列,48条(50%)含有微卫星位点,其中完美型占66%,非完美型22%,混合型12%。微卫星重复基元中,二核苷酸(AG)n和三核苷酸(CAT)n最为常见。The DNA fractions containing microsatellite sequences were captured by hybridizating the digested Rehmannia glutinosa genomic DNA fragments with the oligonucleotide probes （AC）s, （AG）s and （ATG）12 attached to striptavadin coated magnetic beads （Dynal）. The enriched DNA fragments were ligated into pMD 18-T easy vector and then transformed into competent Trans 5 ctcell to form an enriched microsatellite sequence library. PCR screening using M 13 Forward and M 13 Reverse as primers identified 96 clones containing 400 - 800 bp inserts. Sequence analysis of these positive clones confirmed 48 microsatellite sequences, with an enrichment efficiency of 50%. Of these microsatellite loci ,66% were perfect repeat motif,22% imperfect and 12 compound repeat motifs. （AC）n and （CAT）n was the most common motif.

关 键 词：地黄 微卫星 磁珠 富集文库 

分 类 号：Q949.9[生物学—植物学]