地衣型真菌皮革肾岛衣中1个Ⅲ型聚酮合酶基因的克隆与鉴定

Identification and Cloning of a Polyketide Synthase Ⅲ Gene in the Lichenized-fungi Nephrmopsis pallescens

该研究通过对皮革肾岛衣地衣型真菌转录组数据的分析,利用RT-PCR技术,首次克隆得到1个Ⅲ型聚酮合酶基因(NpPKS3),并分析该基因在不同培养基上的表达,以选择皮革肾岛衣地衣型真菌的最佳培养基,为研究NpPKS3基因功能奠定基础,并为异源表达皮革肾岛衣地衣型真菌的聚酮类化合物提供研究材料。结果表明:(1)NpPKS3基因(GenBank登录号MF351559)全长1 335bp,编码444个氨基酸,其产物为NpPKS3蛋白,在细胞质基质中发挥功能。(2)系统进化分析显示,NpPKS3基因编码的氨基酸序列与Ⅲ型聚酮合酶的csyB亚组聚为一支,推测该基因编码csyB类酶蛋白。(3)采用"一菌多产物"策略研究发现,NpPKS3基因在BMG培养基中表达能力最强,在BD培养基上表达能力最弱。

The present study obtained a PKSⅢgene from the transcriptome of the lichenized-fungi Nephrmopsis pallescens.We first cloned full-length cDNA of a PKSⅢgene through the method of RT-PCR,and named it NpPKS3,then we had a trial of NpPKS3gene expression in the different media,to acquire the optimal media culturing the lichen-forming fungi N.pallescens,apply some basis to further explore the function of NpPKS3gene,and simultaneously offer some necessary material to the heterologous expression of polyketide compounds in the lichenized-fungi N.pallescens.The results showed:(1)the full length of NpPKS3gene(GenBank ID:MF351559)has1335bp,codes444amino acid,its transcriptional product could be polyketide synthase of N.pallescens(NpPKS3),and indwells cytoplasmic matrix.(2)The phylogenetic analysis revealed that the NpPKS3amino acids sequence and the csyB of polyketide synthaseⅢclustered into one clade,therefore we could speculate it might be csyB.(3)We detected that the conditions of the NpPKS3gene expression in the different media by OSMAC(one strain,many compounds)strategy.The result showed it is the best expression ability of NpPKS3gene in BMG medium,and the weakest in BD medium.