蛹虫草菌株的ITS序列及系统发育分析

Analysis on ITS Sequence and Phylogeny of Cordyceps militaris Strain

从吉林和辽宁地区采集的野生蛹虫草中分离出10株蛹虫草菌株,提取基因组后扩增ITS序列,与GenBank数据库下载的不同地域来源蛹虫草菌株ITS序列进行碱基组成和变异位点分析;同时对分离株及其继代培养的菌株ITS序列进行碱基变异分析;并从GenBank下载虫草属其他12个种的ITS序列,构建虫草属系统发育树。结果显示:不同地域来源的蛹虫草菌株ITS序列共有5 bp长度变化,碱基C、G含量高于T、A含量;变异位点共有13处,转换发生在T和C之间,转换位点1处,无颠换位点;蛹虫草菌种内的遗传距离在0~0.019之间,虫草属不同物种间的遗传距离为0~0.272;2株继代培养菌株的ITS序列发生碱基突变,原始株与继代培养的菌株的遗传距离分别为0.008和0.005;基于ITS序列构建的虫草属系统发育树显示,蛹虫草菌与九州虫草菌、球孢虫草菌、布氏虫草菌的亲缘关系较近。研究结果表明不同地域的蛹虫草菌株在进化过程中存在差异,同时明确了蛹虫草菌的分类地位及亲缘关系。

Ten Cordyceps militaris strains were isolated from the wild aweto collected in Jilin and Liaoning provinces.Their ITS sequences were amplified after genome extraction,and then base composition and variable site were analyzed between ITS sequences of isolated strains and those of Cordyceps militaris strains from different regions downloaded from GenBank.Meanwhile,analysis on base variation was performed on ITS sequences of isolated strains and their subcultured strains.Furthermore,ITS sequences of other 12 Cordyceps species were downloaded from GenBank to construct phylogenetic tree of Cordyceps.The results showed that ITS sequence of Cordyceps militaris strains from different regions had length variation of 5 bp,and the contents of base C and G were higher than those of base T and A.There were 13 variable sites in ITS sequence,and the transition occurred between T and C with 1 transformation site and no transversional site.Genetic distance of intra-species ranges from 0 to 0.019,and genetic distance of inter-species ranges from 0 to 0.272.ITS sequence of 2 subcultured strains had base mutations,and genetic distances between original strain and subcultured strain was 0.008 and 0.005.Phylogenetic tree bases on ITS sequence of Cordyceps species showed that Cordyceps militaris was closely related to Cordyceps kyushuensis,Cordyceps bassiana and Cordyceps brongniartii.All above results indicated that Cordyceps militaris strains in different regions have differences in the evolutionary process,and taxonomic status and genetic relationship of Cordyceps militaris were also clarified in this study.