牙鲆酵母双杂交cDNA文库的构建及文库质量鉴定  

Construction and identification of a Paralichthys olivaceus yeast two-hybrid library

作　　者：杨长庚[1,2] 刘姗姗[2] 孙冰[2] 陈松林[2] 

机构地区：[1]中国海洋大学海洋生命学院,山东青岛266003 [2]农业部海洋渔业可持续发展综合性重点实验室,中国水产科学研究院黄海水产研究所,山东青岛266071

出　　处：《中国水产科学》2011年第6期1250-1258,共9页Journal of Fishery Sciences of China

基　　金：国家973计划资助项目(2010CB126303);公益性农业行业科研专项(200903046);山东省泰山学者工程专项

摘　　要：为了研究牙鲆(Paralichthys olivaceus)免疫及抗病相关蛋白和大分子物质的功能及相互作用机制,构建了牙鲆酵母双杂交cDNA文库。原始文库滴度为1.6×106 CFU/mL,随机选取单克隆菌落进行PCR扩增,根据PCR扩增结果表明插入片段大部分大于500 bp,平均长度约为1.5 kb,随机调取了115个EST进行了序列测定。扩增后放大文库库容约为1.8×1012 CFU。转化酵母后,文库转化效率为3.6×105CFU/μg,文库库容为5.4×106CFU。牙鲆酵母双杂交文库的构建成功为鲆蝶类免疫机理的研究,尤其是信号分子通路研究提供有效的工具。A yeast two-hybrid cDNA library of flounder （Paralichthys olivaceus） was constructed to investigate the function of immune-related proteins and to gain insight into their mechanisms of interaction. The titer of this original library was 1.6×10^6 CFU/ml. Most of the fragments PCR-amplified from single colonies were larger than 500 bp, with an average length of 1.5 kb. We randomly selected and sequenced 115 expressed sequence tags. The capacity of the amplified library was approximately 1.8×10^12 CFU. After transformation into yeast cells, the transformation efficiency was 3.6×10^5 CFU/μg, and the capacity was 5.4×10^6 CFU. The successful construction of this yeast two-hybrid library provides an effective tool to elucidate mechanisms of immunity, especially signaling pathways, in this flounder species.

关 键 词：牙鲆 酵母双杂交 免疫相关基因 信号通路 

分 类 号：S94[农业科学—水产养殖]