四川凉山地区烟草黑胫病菌的ITS序列分析  

Sequence Analysis of the Ribosomal DNA ITS of Phytophthora Parasitica in Tobacco in Liangshan Region of Sichuan Province

作　　者：张海珊[1] 许大凤[2] 刘东阳[3] 周本国[2] 高正良[2] 卢军[3] 章东方[1] 严丹侃[1] 王芳[2] 王勇[3] 

机构地区：[1]安徽省农业科学院植物保护与农产品质量安全研究所,合肥230031 [2]安徽农业科学院烟草研究所,合肥230031 [3]四川省烟草公司凉山州公司,四川西昌615000

出　　处：《中国烟草科学》2015年第1期96-101,共6页Chinese Tobacco Science

基　　金：四川省烟草公司凉山州公司科技项目"凉山州烟草黑胫病发生规律及防治研究"(201203002)

摘　　要：本研究利用真菌核糖体基因转录间隔区（ITS）通用引物,PCR扩增四川省凉山州不同地区的20株烟草黑胫病菌的ITS序列,并对PCR产物进行了克隆测序。结果表明,20个供试菌株的ITS1-5.8S-ITS2总长均为803 bp;各菌株之间的ITS序列同源性达到99.3%以上;与Gen Bank报道的寄生疫霉台湾烟草分离株Phytophthora parasitica（GU111675.1）的同源性在99.6%-100%之间;但各菌株ITS序列的个别位点存在突变,这些突变主要集中在ITS2区上,其中第468处的突变具有一定地域性,主要为德昌和西昌的菌株。利用MEGA6软件对20个供试菌株及Gen Bank中登陆的15个疫霉属菌株序列进行聚类分析,供试菌株与Gen Bank上登录的GU111675.1、KF010303.1、AJ854295.1、KC768775.1、L41383聚在同一聚类组上,均为Phytophthora parasitica（或Phytophthora nicotianae）,其它疫霉种聚在不同的聚类组上,遗传关系较远,这表明来自四川凉山不同地区的20株烟草黑胫病分离菌株均为寄生疫霉（Phytophthora parasitica）。The internal transcribed spacer regions （ITS） of the ribosomal RNA gene from 20 isolates of tobacco black shank wereamplified with the polymerase chain reaction through the universal primers and sequence. The result showed that the total length ofITS of those 20 isolates tested were all 803bp, and the nucleotide sequence identity among those sequences was above 99.3%, and 20strains had 99.6%-100% homologies with Phytophthora parasitica（GU111675.1） published in GenBank. But the ITS of those 20isolates tested had some mutation sites which mainly concentrated in the ITS2 area and 468 mutation of isolates, which mainly camefrom Dechang and Xichang had certain regional. With the bio-software of MEGA6, Sequences from the above isolates werecompared with 15 sequences published in GenBank database and phylogenitic trees were constructed based on ITS sequence data.The results showed that the isolates tested, GU111675.1, KF010303.1, AJ854295.1, KC768775.1 and L41383 were clustered into thesame clade and all were Phytophthora parasitica （or Phytophthora nicotianae）. They were clustered in a distant clade for differentPhytophthora species and phylogenetically distinct each other. The results indicated that twenty isolates which isolated from differentregions of Liangshan prefecture were Phytophthora parasitica.

关 键 词：烟草黑胫病 ITS 寄生疫霉 烟草 

分 类 号：S435.72[农业科学—农业昆虫与害虫防治]