目的 克隆药用真菌猪苓Polyporus umbellatus凋亡抑制因子基因BI-1并进行生物信息学和表达模式分析。方法 利用5'-RACE PCR方法获取基因cDNA全长;利用生物信息学软件预测蛋白的理化性质、结构域和跨膜结构等分子特性;采用Bioeditor以及MEGA 6.0分别进行氨基酸多序列比对和进化关系分析;实时荧光定量PCR分析基因表达模式。结果 猪苓细胞凋亡抑制因子BI-1(Genebank登录号JQ693683)的cDNA全长为1 091 bp,其中编码区占1 005 bp,编码334个氨基酸,推测相对分子质量为36 170,理论等电点为10.51,定名为PuBI-1。整个多肽链具5个疏水区,表现为疏水性,是疏水性蛋白。系统进化树结果显示PuBI-1所在分支隶属于担子菌群,与裂褶菌形成一单系。PuBI-1基因转录本在初始期和生长期表达量在菌核组织中显著高于未形成菌核的菌丝组织,分别为原基期菌核(SI)为3.8倍、发育期菌核(SD)为7.497倍,成熟期菌核和菌丝中的PuBI-1基因的表达量几乎无差异。结论 猪苓凋亡抑制因子基因BI-1的分子特征及表达模式为进一步研究其在猪苓菌丝形成菌核过程中的作用奠定理论基础。

Objective To clone the Bax inhibitor-1 (BI-1) gene from Polyporus umbellatus and perform the bioinformatics and expression mode analysis. Methods To clone the full-length cDNA of BI-1 using RACE technology. The characteristics of physiochemical properties, conserved domains, and transmembrane domain of the predicted BI-1 protein were determined using bioinformatic tools. Results The full-length cDNA of BI-1 gene was 1 091 bp in length and encoded a 334-aa protein with a molecular weight of 36 170 and an isoelectric point (pI) of 10.51. The polypeptide chain was a hydrophobin with five hydrophobic regions. The PuBI-1 belonged to basidiomycete group according to the phylogenetic analysis. Real time quantitative PCR (RT-qPCR) analysis revealed that the transcription level of PuBI-1 gene was significantly higher in the beginning and developing stages of sclerotial formation with 3.8 and 7.497 fold over those in the mycelium, but the transcripts decreased sharply with the sclerotial development. Conclusion Molecular characterization and expression patten of PuBI-1 gene will be useful for the further functional determination of the gene during the development of P. umbellatus sclerotium.

国家自然科学基金资助项目(31201666);设施食用菌高效碳循环研究与示范(FT2014-03)