链格孢属真菌的分子复核鉴定及系统发育研究

Molecular Identification and Phylogeny of Genus Alternaria

为了验证中国林业微生物保藏管理中心库藏链格孢属(Alternaria Nees)菌种资源分类地位的准确性,探索一种链格孢属真菌的快速复核鉴定方法,采用ITS序列分析、NCBI blast以及系统发育分析等方法,对中国林业微生物保藏管理中心库藏链格孢属的36个种及未定种的375个菌株进行分子复核鉴定以及系统发育关系研究。结果表明:研究的375个菌株中,有21个菌株的分类地位不准确,其中有9株菌的ITS序列与链格孢属菌种的同源性在92%～96%之间,另外12株菌的ITS序列与链格孢属菌种的同源性在60%～85%之间。基于ITS序列构建的NJ树显示,29个小孢子种的37个生物型形成了一个稳定的分支,而6个大孢子种与外群Ulocladium sp.聚在一个分支。本研究中,ITS序列分析可以用来区分和鉴定形态差别明显的大孢子种,但无法区分亲缘关系较近的29个小孢子种。

In order to identify cultures of Alternaria Nees from China forestry culture collection center at molecular level by ITS sequence analysis. The internal transcribed spacer （ITS） region DNA fragment of 375 strains were amplified and sequenced by universal primers ITS1 and ITS4. And all the sequences of the 375 strains in present study were compared with the ITS sequences of related Alternaria species available in the GenBank database by BLAST search. Phylogenetic analysis was performed using the neighbor-joining method based on Paup 4.0 bl0. ITS sequencing and NCBI BLAST results showed that： classification status of 21 strains from 375 trains were wrong, in which 9 strains had 92%-96% similarities with the reference strains, and the other 12 strains had 60%-85% similarities with the reference strains. The phylogenetic tree showed that 29 species were formed one cluster with 100% boot strap support, and the other seven species were divided into 6 clusters. The results indicated that： the ITS sequence could not be used to the separation of 29 species of Alternaria, but the other 6 species could be differentiated each other.