中国被毛孢分离鉴定及其<i>PKS、NRPS</i>功能基因的多样性

任朝辉; 牛晓娟; 杜静; 张传博

doi:10.13386/j.issn1002-0306.2018.18.022

中国被毛孢分离鉴定及其PKS、NRPS功能基因的多样性

Isolation and Identification of Ophiocordyceps sinensis and the Diversity of Its' PKS and NRPS Functional Genes

摘要

摘要: 探究中国被毛孢(Ophiocordyceps sinensis)遗传、聚酮合成酶(Polyketide synthase,PKS)、非核糖体多肽合成酶(Non-ribosomal peptide synthase,NRPS)的多样性,为寻找合成生物活性物质的潜在菌株奠定基础。采用组织分离法从冬虫夏草中分离获得一株具有较强抑菌活性的真菌ZG4-23,采用多基因系统分析的方法对该菌株进行鉴定;利用兼并性引物对菌株ZG4-23的PKS基因和NRPS基因进行PCR扩增及序列测定分析,构建系统发育树,明确菌株ZG4-23的PKS基因序列和NRPS基因序列的系统进化地位。结果表明,多基因序列分析显示,菌株ZG4-23与多个中国被毛孢菌株同源性高达99%,因此确定该菌株为中国被毛孢(O. sinensis)。菌株基因组含有PKS I、NRPS 基因,经比对,PKS I片段与已知Ophiocordyceps sinensis PKS序列相似度为99%,NRPS片段与Trichophyton rubrum NRPS序列相似度为76%。中国被毛孢具有较强合成生物活性次生代谢产物的潜在能力,值得进一步开发研究和应用。

Abstract: To explore the inheritance of Ophiocordyceps sinensis and the diversity of the polyketide synthase (PKS) and non-ribosomal peptide synthase (NRPS) genes and to lay the foundation of looking for the potential strains of the synthesis of bioactive substances. A strain of fungi ZG4-23 with strong antibacterial activity was isolated from O. sinensis by tissue isolation method. The multilocus phylogenetic analysis was used for identification of the fungi. PKS and NRPS genes fragments were PCR amplified from genomic DNA with degenerate primers. The biosynthetic potential of strain ZG4-23 was accessed by bioinformatic and phylogenetic analyses of PKS and NRPS genes sequences. The polygenes sequence analysis showed that, the sequence homology was 99% between the strain ZG4-23 and the multiple strains of O. sinensis.The strain contained PKS I gene and NRPS gene. The PKS I sequence had 99% homology compared with O. sinensis PKS gene. The NRPS gene sequence had 76% homology with Trichophyton rubrum NRPS gene. This study demonstrated that O. sinensis had potential capacity to synthesize bioactive secondary metabolite, which was worth of further research and development.