勒氏笛鲷微卫星位点的筛选及特征分析.pdf

HEREDITAS (Beijing) 2007 3 , 29(3): 355―359 ISSN 0253-9772 研究报告 DOI: 10.1360/yc-007-0355 1 2 1,2 1 郭昱嵩 , 王中铎 , 刘楚吾 , 刘筠 1. , 410081; 2. , 524025 : 采用PCR法快速筛选勒氏笛鲷(Lutjanus russelli)基因组文库, 以获得(CA) 微卫星位点。勒氏笛鲷基因组 n DNA经限制性内切酶HaeⅢ+ Dra Ⅰ双酶切后, 连接T-载体克隆, 构建基因组文库。以通用引物M13+/- 与重复序 列引物(CA)15对基因组文库进行筛选, 二次筛选后得到121 个可能含有微卫星位点的阳性克隆。进行序列测定, 共获得53 个CA(n ≥7)重复序列, 重复次数主要分布于7~15(80.77%) 。在所得微卫星序列中, 重复单元除CA外, 还观察到单碱基、三碱基、四碱基、五碱基重复单元。根据侧翼序列设计48 对引物, 通过优化PCR反应条件, 可 获得清晰可重复的目的条带。研究旨在为勒氏笛鲷遗传多样性研究及遗传图谱的构建等奠定基础, 为勒氏笛鲷 资源的合理开发利用提供参考。 : 微卫星; 勒氏笛鲷; PCR Rapid isolation and characteristics analysis of microsatellites from Lutjanus russelli 1 2 1,2 1 GUO Yu-Song , WANG Zhong-Duo , LIU Chu-Wu , LIU Yun 1. College of Life Science, Hunan Normal University , Changsha 410081, China; 2. Fisheries College, Guangdong Ocean University, Zhanjiang 524025, China Abstract: The (CA) DNA sequences in Lutjanus russelli were isolated through PCR arrays, and the characteristics of the n microsatellite were analyzed. DNA was extracted from a sample of Lutjanus russelli, and digested with Hae+Dra . The fragments were ligated to pUCm-T vector and transferred into DH5α to construct a genomic library. The positive clones were isolated with universal M13 reverse and forward primers, M13 forward primer and the simple sequence repeats (SSRs) probes (CA)15,