应用反向斑点杂交技术从转录序列的选择性捕获中筛选副猪嗜血杆菌表达差异基因  

Screening Differentially-expressed Genes of Haemophilus parasuis from the Selective Capture of Transcribed Sequences (SCOTS) by Reverse Dot-blot Hybridization

作　　者：胡军勇[1] 金卉[1] 谢启运 万云[1] 楚杰[1] 刘翠云[1] 陈焕春[1] 周锐[1] 

机构地区：[1]华中农业大学动物医学院,农业微生物学国家重点实验室,武汉430070

出　　处：《农业生物技术学报》2009年第2期189-194,共6页Journal of Agricultural Biotechnology

基　　金：高等学校科技创新工程重大项目培育资金项目(No.706042);国家高技术研究与发展计划(No.2007AA02Z193);国家科技支撑计划(No.2006BAD06A01);湖北省自然科学基金青年杰出人才项目(No.2007ABB015)资助

摘　　要：通过对靶DNA的种类和浓度、探针浓度、紫外交联时间等反向斑点杂交条件优化,建立了一种从选择性捕获的转录序列(selective capture of transcribed sequences,SCOTS)库中筛选副猪嗜血杆菌在体内、外差异表达的基因的方法。通过反向斑点杂交技术从10个样品中筛选出的6个样品被证明是副猪嗜血杆菌(Haemophilus parasuis)在体内上调表达或者是特异表达的基因。结果表明:SCOTS技术能够有效地获得一个可能存在差异表达基因的cDNA文库,而反向斑点杂交技术可以有效地从中筛选到差异表达基因。A reverse Dot-blot hybridization （RDH） method was established for screening the differentially-expressed genes of Haemophilus parasuis in vivo and in vitro from the selective capture of transcribed sequences （SCOTS） pools by optimizing the hybridization conditions including the kinds and concentration of target DNA, probe concentration, and UV irradiation time. Six samples were verified to be differentially-expressed genes in vivo of H. parasuis from ten samples of cDNA libraries （SCOTS pools） by RDH. The results showed that SCOTS was an effective approach to establish cDNA libraries containing differentially-expressed genes and RDH could be used to effectively pick up the differentially-expressed genes from the SCOTS pools.

关 键 词：副猪嗜血杆菌 差异表达基因 转录序列的选择性捕获 反向斑点杂交 

分 类 号：S188[农业科学—农业基础科学]