文献类型：期刊文献

中文题名：应用抑制差减杂交法分离粗枝大叶黄杨幼苗的冷诱导表达基因

英文题名：Isolating Cold-Regulated Genes from Euonymus japonicus ‘Zhuzi’ Seedlings through Suppression Subtractive Hybridization

作者：孙洪波[1] 王国英[2] 孙振元[3] 古润泽[4] 赵梁军[1]

第一作者：孙洪波

机构：[1]中国农业大学观赏园艺与园林系;[2]中国农业大学农业生物技术国家重点实验室;[3]中国林业科学研究院花卉研究中心;[4]北京市园林科学研究所

年份：2005

卷号：38

期号：1

起止页码：135-139

中文期刊名：中国农业科学

外文期刊名：Scientia Agricultura Sinica

收录：CSTPCD;;Scopus;北大核心:【北大核心2004】；CSCD:【CSCD2011_2012】；

基金：北京市科委"抗旱;耐寒转基因地被植物的培育"资助项目(H020620110130)

语种：中文

中文关键词：幼苗;抑制差减杂交;大叶黄杨;低温处理;同源序列;低温诱导;技术分离;基因;DNA;诱导表达

外文关键词：Euonymus japonicus ‘Zhuzi’; Cold-induced gene; Suppression subtractive hybridization (SSH); cDNA macroarray

分类号：S511.9;R644

摘要：应用抑制差减杂交(suppressionsubtractivehybridization,SSH)方法,构建冷诱导表达的正向抑制差减cDNA文库,低温处理的幼苗为tester,常温处理为driver,通过cDNA宏阵列差异筛选cDNA文库,得到604个低温诱导或表达增强的候选克隆,对其中的84克隆进行DNA测序,去除冗余的cDNA,在GenBank中进行核酸和蛋白质同源性的比较和功能分析,共有36个单一序列,有12个cDNA未找到同源序列,可能为新基因,表明采用抑制差减杂交方法与cDNA宏阵列技术分离诱导表达的基因是可行的。
Suppression subtractive hybridization (SSH) was utilized for the isolation of cDNA fragments for Euonymus japonicus ‘Zhuzi’ differentially expressed genes, and forward suppression subtractive cDNA Library of cold-regulated gene was constructed. The seedlings of Euonymus japonicus ‘Zhuzi’ were treated with low temperature as tester and untreated seedlings as driver. Subtractive cDNA Library was differently screened through cDNA macroarray, six hundreds and four cDNA clones were identified as cold specifically induced or highly expressed. After sequencing of 84 cDNA clones and removing redundant cDNAs, 36 cold-regulated unique cDNA clones were obtained. The results showed that 12 cDNA clones are expected to be novel genes, because no sequence homology with any known sequences was found in GenBank databases. This indicated that it is possible to use SSH and cDNA macroarray in the isolation of cold-regulated genes.