绵羊肺炎支原体特异性分子检测靶标的筛选与应用  

Screening and application of specific molecular detection targets for Mycoplasma ovipneumoniae

作　　者：陈芙 储岳峰[2] 刘保红[2] 陈胜利[2] 郝华芳[2] 王鹏雁[1] 颜新敏[2] CHEN Fu;CHU Yue-feng;LIU Bao-hong;CHEN Sheng-li;HAO Hua-fang;WANG Peng-yan;YAN Xin-min(College cf Animal Science and Technology,Shihezi University,Shihezi 832001,China;State Key Laboratory of Veterinary Etiological Biology,Lanzhou Veterinary Research Institute,Chinese Academy qf Agricultural Sciences.Lanzhou 730046,China)

机构地区：[1]石河子大学动物科技学院,新疆石河子832001 [2]中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室,甘肃兰州730046

出　　处：《中国兽医科学》2021年第7期848-856,共9页Chinese Veterinary Science

基　　金：国家重点研发计划项目(2016YFD0500907);中国农业科学院兰州兽医研究所“元亨人才”计划项目(农科兰兽2020-119)。

摘　　要：本文旨在筛选出特异性的绵羊肺炎支原体(Mycoplasma ovipneumoniae)分子检测靶标,利用全基因组BLASTn筛选出绵羊肺炎支原体特异性蛋白基因序列,再运用NCBI线上Primer BLAST程序,选择G/C含量较高的4个靶基因设计11对引物,通过引物筛选和反应条件的优化,成功筛选到一对针对靶标728序列的特异性引物728 2F/2R。用该引物对53株绵羊肺炎支原体菌株、15株其它支原体和18株常见反刍动物细菌的DNA进行扩增,并对27份临床样品进行检测。结果显示,7282F/2R只能从绵羊肺炎支原体菌株DNA扩增出288 bp目的片段,且具有良好的敏感性,与已发表方法相比特异性更好,更适合于绵羊肺炎支原体检测。The aim of this study is to screen a new molecular detection target of Mycoplasma ovipneumoniae(Mo).The specific protein-coding gene sequences of Mo were selected by the whole genome BLASTn,and then 11 pairs of primers were designed by using the Primer BLAST program of NCBI online.Through primers screening and optimization of reaction conditions,a pair of specific primer 728 2F/2R targeted the gene 728,was successfully selected.The specificity of the primers was assessed with following DNA samples,53 strains of Mycoplasma ovipneumoniae,15 strains of other mycoplasmas,18 strains of bacteria commonly from ruminants and 27 clinic samples.The results showed that the PCR with the primers of 728 2F/2R can amplify a 288 bp fragment from M.ovipneumoniae strains only,and with high sensitivity.It has been proved that the PCR established here has better specificity than that of the published PCR and it is more suitable for M.ovipneumoniae detection.

关 键 词：绵羊肺炎支原体 分子诊断靶标 筛选 

分 类 号：S852.62[农业科学—基础兽医学]