PJ综合症相关抑癌基因LKB1/STK11的逆转录PCR法克隆及序列分析

Cloning and Sequencing of Tumor Suppressor Gene LKB1/STK11 Associated with Peutz-Jeghers Syndrome by RT-PCR

从正常人胎盘组织中克隆出人野生型PJ综合症相关抑癌基因LKB1/STK11cDNA片段.以提取的总RNA为模板,采用逆转录法获得cDNA第一条链,采用94℃变性、72℃退火及延伸的特殊条件,经PCR扩增出抑癌基因LKB1/STK11cDNA片段,克隆到T载体,在国内首次报道了LKB1/STK11基因的克隆.序列分析表明,该cDNA全长1299bp,与GenBank中人野生型LKB1cDNA编码序列完全相同,证实获得了人野生型抑癌基因LKB1/STK11的cDNA克隆.LKB1/STK11cDNA的成功克隆为其原核表达载体的构建和LKB1/STK11蛋白在细胞内的作用及其抑癌效果、抑癌机理的研究打下了基础.

The tumor suppressor gene LKB1 / STK11 associated with Peutz-Jeghers syndrome was cloned from human placenta tissues for the first time in China. The total RNA was isolated from human placenta tissues. The first strand of cDNA was obtained by reversed transcription and the cDNA fragment of LKB1 / STK11 was amplified by optimized PCR under the following cyclic conditions： denaturing at 94 ℃, annealing and extension at 72 ℃. This fragment was cloned into the vector pMD18-T, and then introduced into E. coli DH5a. The sequence analysis showed that cDNA obtained was 1299bp in length and was proved to be identical with the reported LKB1 / STK11 cDNA sequence in GenBank. The successful cloning of LKB1 / STK11 gene provides the basis for the construction of its prokaryotic expression vector and further study on the tumor suppression and other functions of LKB1 / STK11 protein in cells.