球孢白僵菌DNA甲基转移酶基因Dim-2的克隆及其表达分析

Cloning and Expression Analysis of DNA Methyltransferases Encoding Gene Dim-2 from Beauveria bassiana

本研究运用RACE技术,从球孢白僵菌中克隆出完整的DNA甲基转移酶基因Dim-2的编码区序列。该基因c DNA全长4021 bp,5′端非翻译区283 bp,3′端非翻译区303 bp,开放阅读框(ORF)3429 bp,编码1142个氨基酸。蛋白理论分子量为128 k D,理论等电点为6.13。结构域分析显示,该基因编码蛋白含有1个BAH结构域和合成5-甲基胞嘧啶的活性区域。Real-time PCR与HPLC检测表明,球孢白僵菌DNA甲基化程度会随着其生长发育的变化而不断改变。结果显示,球孢白僵菌Dim-2基因的转录表达量与基因组DNA的5-甲基胞嘧啶百分含量在菌体培养初始产孢阶段(即培养第7 d),均出现了1个低谷。这可能意味着球孢白僵菌Dim-2基因所引起的DNA甲基化与其生长发育之间具有内在的联系。本研究将为进一步探索DNA甲基化在虫生真菌生长发育中具体功能机制奠定基础。

The full-length c DNA of Dim-2 gene was cloned from Beauveria bassiana using a RACE technique. The c DNA of Dim-2 had a lenght of 4021 bp, including an open reading frame(ORF) with 3429 bp encoding 1142 amino acids. The protein had a molecular mass of 128 k D with a calculated p I of 6.13. The Realtime-PCR and HPLC analysis indicated that the degree of DNA methylation varied along with fungal growth development. The results showed that both the expression level of Dim-2 gene and the degree of DNA methylation reached the minimum levels at initial stage of sporulation(the 7th day). It was inferred that there was an internal relationship between DNA methylation caused by Dim-2 gene and growth development in B. bassiana. The study will benefit future functional studies of DNA methylation in developmental control of entomopathogenic fungi.