重伞灵芝线粒体基因组在灵芝种间鉴定中的应用

Application of the Ganoderma multipileum mitogenome to identification of Ganoderma species

线粒体是"动力工厂",能够进行氧化代谢实现能量的转化,参与三羧酸循环形成ATP。随着分子生物学和生物信息学技术的发展,担子菌灵芝属中已有几种灵芝的线粒体基因组被相继报道,但尚未有对重伞灵芝线粒体基因组的报道。本研究通过对重伞灵芝YX线粒体基因组进行组装注释,比较分析了与其他灵芝属物种的差异,根据差异序列构建分子标记对重伞灵芝菌株快速鉴定。结果显示重伞灵芝线粒体基因组大小为67340bp的闭合环形结构,含有15个普通蛋白编码基因,28个tRNA基因,以及rRNA的大小亚基基因。共5个基因含有12个内含子,主要为IB型,部分为ID型,包含LAGLIDADG1superfamily、GIY-YIGCterm superfamily保守结构域。根据重伞灵芝线粒体cox2基因设计的特异性引物扩增结果显示:4株重伞灵芝的cox2基因片段均可准确扩增,且扩增不出其他灵芝物种的cox2基因片段。基于各灵芝菌株线粒体cox2基因序列构建系统进化树,可以将4株重伞灵芝归在同一分支上,并且同源性为98%,与ITS鉴定结果一致。可见,基于线粒体基因cox2特异性引物可以快速地对重伞灵芝进行鉴定,且只需通过观察扩增条带的有无,无需测序,省时省力。

Mitochondria are "power plants" of the cell that are capable of oxidative metabolism for energy conversion, participating in the tricarboxylic acid cycle to form ATP. With the development of molecular biology and bioinformatics technology, several mitochondrial genomes of Ganoderma species have been reported recently. However, there has been no report on the mitochondrial genome of G. multipileum. In this study, the mitochondrial genome of G. multipileum YX was assembled and annotated, and the differences between the mitogenome and that of other Ganoderma species were compared and analyzed. According to differential sequences, the molecular markers were constructed. It is indicated that the mitogenome of G. multipileum has a closed loop structure of 67 340 bp, containing 15 common protein coding genes, 28 tRNA genes, and subunit gene of rRNA. There are five genes in total, containing 12 introns which are of mainly IB type and partly ID type and including LAGLIDADG1 superfamily, and GIY-YIGCterm superfamily conserved domain. According to the specific primers designed by the mitochondrial cox2 gene of G. multipileum, the cox2 gene fragments of four G. multipileum strains can be accurately amplified while the cox2 gene fragments of other Ganoderma species cannot be amplified. Based on the systematic evolutionary tree constructed by mitochondrial cox2 gene sequences of G. multipileum strains, four strains of G. multipileum can be classified into the same branch, with homology of 98%, being consistent with the ITS identification results. The result proves that G. multipileum can be quickly identified by mitochondrial gene-based cox2-specific primer, and we only need to observe the presence or absence of the amplified band without sequencing.