白念珠菌角鲨烯环氧化酶基因开放读框的克隆及其在大肠杆菌中的表达

MOLECULAR CLONING AND EXPRESSION OF SQUALENE EPOXIDASE FROM CANDIDA ALBICANS IN E.COLI

根据白念珠菌角鲨烯环氧化酶基因的开放读框中编码1MSSVKY6的序列和编码492NEIVR496的序列分别设计上、下游引物,以白念珠菌ATCC11006的基因组DNA为模板进行PCR扩增;将PCR产物克隆并做序列分析后,在大肠杆菌中进行表达。结果表明PCR获得大小约为1.5kb的产物,测序分析表明克隆的产物大小为1491bp,正是白念珠菌角鲨烯环氧化酶基因的开放读框,表达得到约为80kDa大小的蛋白,与理论计算一致。本研究为开展特比萘芬与其作用靶酶关系的研究奠定了基础。

The primers, which contain the sequences corresponding to amino acid 1MSSVKY6 and 492NEIVR496 of squalene epoxidase from Candida albicans, were synthesized. Polymerase chain reaction (PCR) was carried out using these primers and genomic DNA from C.albicans ATCC11006 as template. The PCR product was expressed in E. coli. after being cloned and sequenced. The results showed that the PCR product is 1491bp , which is just the open reading frame (ORF) of squalene epoxidase gene from Candida albicans. A protein about 80kDa was generated after the cloned DNA fragment was expressed in E. coli. This study provided the basis for the further investigation on the interaction between terbinafine and its target enzyme.