柞蚕微孢子虫Nosema pernyi微管蛋白基因的克隆及系统发育分析

【目的】柞蚕微粒子病的病原为柞蚕微孢子虫Nosemapernyi,为解明柞蚕微孢子虫微管蛋白基因的序列信息,明确柞蚕微孢子虫的系统分类学地位。【方法】采用RT-.PCR、3′RACE(Rapid amplification ofcDNAends)等技术克隆得到了柞蚕微孢子虫的α、β和y-微管蛋白基因,并利用α、β-微管蛋白序列,分别采用NJ、ML法构建进化树。【结果】将克隆得到的基因序列提交NCBI(GenBank登录号:KF154086、KF023271、KF740389)。构建的系统发育树显示,微孢子虫类以一个独立群位于真菌群体中,与真菌的虫霉门关系较近,且与担子菌、球囊菌、壶菌、接合菌及部分子囊菌互为姐妹群。从部分微孢子虫的系统发育分析结果可以看出,20种微孢子虫分为2个分支,柞蚕微孢子虫与其他Nosema属聚为一类。【结论】本研究克隆得到了柞蚕微孢子虫α、β和y-微管蛋白基因,系统发育分析为更进一步了解柞蚕微孢子虫奠定了基础。

Cloning and phylogenetic analysis of the tubulin genes of Nosema pernyi

Objectives] The purpose of this project was to investigate tubulin gene sequences of Nosema pernyi in order to provide a foundation for further phylogenetic analysis. Methods] Using RT-PCR, 3＇ RACE technology, α, β and γ-tubulin genes of N. pernyi were cloned. Neighbour-Joining and Maximum Likelihood were used to construct a phylogenetic tree. Results] a, fl and ）,-tubulin eDNA sequences of N. pernyi were submitted to NCBI （GenBank accession numbers： KF154086, KF023271, KF740389）. Phylogenetie analysis shows that the microsporidia are a single group in the fungi that are most closely related to the Entomophthoromycota, and are sistergroups to the Basidiomycota, Chytridiomyeota, Zygomyeota, Glomeromycota and some of the Aseomyeota. N. pernyi, together with other Nosema spp., belongs to one of two groups in the microsporidia. Conelusion] α, β and γ-tubulin genes ofN. pernyi can be successfully cloned and the resultant phylogenetie analysis provides a foundation for further taxonomic research on N. pernyi.