艰难梭菌细胞毒素B功能区的克隆及序列分析

Cloning and sequencing of the C-terminal domain gene of Clostridium difficile toxin B

目的克隆艰难梭菌(Clostridium difficile,C.d)细胞毒素B羧基末端功能区(CDB3)基因,并对其进行测序及生物信息学分析。方法利用PCR技术扩增CDB3基因,并将其定向插入pET-22b(+)载体中,以DNA自动分析仪进行序列测定,并以生物信息学软件分析其生物学特性。结果成功克隆了艰难梭菌CDB3基因,经测序表明与GenBank中分布的Clostridium difficile VPI10463的ToxinB3基因序列完全一致。DNAstar软件预测其蛋白质的相对分子量(Mr)约为71.3 kD,并显示出良好的抗原性。结论研究获得了序列正确的CDB3基因,为其重组表达及其相关研究奠定了良好基础。

Objective To clone the C-terminal domain gene （CDB3） of Clostridium difficile （C. d） and to perform sequencing and analyse, the biological information. Methods The CDB3 DNA was amplified by PCR. The PCR products were insered directionally into vector pET-22b （ ＋ ） to construct recombinant danes of CDB3 and was sequenced. The biological property at the amiono acid level was analyzed by DNAstar 5.0. Results The recombinant plasmid was constructed. DNA sequence analysis showed the sequence of CDB3 was the same as what was published by GenBank. DNAstar 5.0 software predicted its relative molecular mass （Mr） was 71.3 kD and possessed good antigenieity. Conclusion A confirmed CDB3 gene has been obtained, providing a good foundation for recombination, expression and related study.