不同含油量大豆成熟期胚抑制消减cDNA文库构建及其差异表达基因分析  

Construction of suppression subtractive cDNA library and analysis of differentially expressed genes during maturing period seed of high and low oil content soybean variety

作　　者：程金朋[1] 陈波[2] 徐有明[1] 周新安[2] 卢长明[2] 魏文辉[2] 

机构地区：[1]华中农业大学园艺林学学院,湖北武汉430070 [2]中国农业科学院油料作物研究所基因组与分子生物学研究室,湖北武汉430062

出　　处：《中国油料作物学报》2007年第4期365-371,共7页Chinese Journal of Oil Crop Sciences

基　　金：中国农业科学院油料作物研究所所长基金(200602)

摘　　要：利用抑制性消减杂交(suppression subtractive hybridization,SSH)技术,以高含油量大豆品种中豆32和低含油量大豆品种油春02-6受精后36d胚为实验材料构建消减cDNA文库。文库的插入片段集中在500bp左右,挑取766个克隆进行PCR筛选获得700个阳性克隆,经过点杂交筛选后选择了575个阳性克隆进行测序。经过同源性比对归并后得到237个差异表达基因,除10个未知基因外,其余涉及到蛋白贮藏、蛋白质降解、激素调节等多个方面。本文还对部分可能与油脂合成相关的差异表达基因进行了RT-PCR半定量和荧光定量PCR检测,并简要讨论了它们的功能,为进一步研究这些基因在大豆油脂合成过程中的作用提供依据。The 36 clay after ferfilization（DAF） embryoes of high oil content variety Zhongdou 32 and low oil content variety Youchun 02-6 were used to construct suppression subtractive eDNA library. The inserted fragment sizes focus on 500 bp,and 766 clones were verified by PCR, among which 700 clones were used to perform clot blot hybridization. 575 clones were obtained and sequenced. 237 differentially expressed genes were obtained by homologous BLAST search. Besides 10 unknown function genes, the others related to protein storing, protein degradation, auxin regulation and so on. Parts of differentially expressed genes probably relating to oil synthesis were detected by RT-PCR and Real-time quantitative PCR,their functions were also primarily discussed,which will be a guidance for studying the functions of these genes during oil synthesis.

关 键 词：大豆 抑制性消减杂交 CDNA文库 RT—PCR 实时荧光定量PCR 

分 类 号：S565.103[农业科学—作物学]