小麦印度腥黑粉菌线粒体DNA提取及其ATP6基因在真菌遗传发育分析中的应用

Extraction of mitochondrial DNA of Tilletia indica Mitra and application of the ATP6 gene on fungal phylogenetic analysis

摘要:

小麦印度腥黑粉菌(Tilletia indica Mitra)是一种世界范围的重要检疫性有害真菌,该病原菌和近似种之间冬孢子的形态特征极为相似,遗传关系非常相近。为了从分子水平上探讨小麦印度腥黑粉菌和近似种之间线粒体基因序列的差异,从新鲜菌丝中提取总DNA,经两次氯化铯密度梯度超速离心分离线粒体DNA(mtDNA),提取的mtDNA纯度较高,可用于克隆、酶切分析和PCR扩增等分析。选取基因ATP(adenosine triphosphate)6的序列,并结合GenBank中相关种类的ATP6基因DNA序列进行了系统发育分析。结果表明,线粒体基因ATP6可用于科属水平的分类鉴定。

Abstract:

Tilletia indica Mitra，an important pest around the world which is the causative agent of Karnal Bunt of wheat，was very close to Tilletia walkeri phylogenetically and morphologically. 69.6ng mitochondrial DNA (mtDNA) of Tilletia indica obtained from 6.4mg total DNA by the method of CsCl/bisbenzimide density gradient ultracentrifugation can be used for electrophoresis，cloning，enzyme restriction analysis and PCR amplification. The ATP6 gene sequence was cloned from the fragment of mtDNA and sequenced for analysis of phylogenetic tree with the other related sequences in GenBank by the software PAUP to reveal the phylogenetic relationships within the Tilletia species.