绵羊肺炎支原体Y98株未知基因Hsp70(DnaK)的克隆  

Using tail-PCR technology to clone the unkonwn gene Hsp70(DnaK) of Mycoplasma ovipneumoniae Y98

作　　者：马春骥[1] 李敏[1] 赵东[1] 王玉炯[1] 

机构地区：[1]宁夏大学西部特色生物资源保护与利用教育部重点实验室,宁夏银川750021

出　　处：《中国兽医学报》2011年第12期1733-1737,1786,共6页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(30860207;30960289);宁夏回族自治区科技攻关计划资助项目(Z2006-1-75001);宁夏高等学校科学技术研究重点项目

摘　　要：绵羊肺炎支原体(Mycoplasma ovipneumoniae,MO)是绵羊传染性胸膜肺炎(Contagious ovine pleuropneumo-nia)的主要致病菌。热休克蛋白Hsp70也叫(DnaK),具有分子伴侣和免疫的作用。本试验以MO Y98基因组为模板,通过比对15种支原体Hsp70基因的序列并设计简并引物,分别利用同源克隆和染色体步移—Tail-PCR技术克隆到了四段Hsp70(DnaK)基因片断并进行了核苷酸序列测定。利用序列拼接软件对克隆的四段序列进行序列拼接,通过ORF Finder软件分析出了MO Y98的Hsp70的开放式阅读框架,预测分析该基因是由1 815bp组成,编码604个氨基酸。本试验于国内外首次克隆到了MO的Hsp70基因序列,为MO的抗原研究以及Hsp70的功能研究等奠定了基础。Mycoplasma ovipneumonia（MO） is the main pathogen causing contagious ovine pleuropneumonia.In this work,comparing 15 species Hsp70 gene and protein sequence of mycoplasma,the conservative sequences were found and degenerate primers were designed.Then,four fragments of MO Hsp70 gene were acquired by homologous cloning combined with Tail-PCR,chromosome walking.Four fragments of MO Hsp70 were assembled and the whole ORF was found by ORF Finder,which was 1815 bp in length that encoded 604 amino acids.MO Hsp70 was cloned the first time at home and abroad.Our results layed the foundation for the further research of Hsp70 function and MO antigens.

关 键 词：绵羊肺炎支原体 HSP70 同源克隆 TAIL-PCR 

分 类 号：S852.62[农业科学—基础兽医学]