干扰素诱导的跨膜蛋白-1基因的扩增、克隆及蛋白表达

目的研究干扰素诱导的跨膜蛋白-1（interferon-inducible transmembrane protein-1,IFITMP-1）基因的扩增、克隆及蛋白表达.方法以含IFITMP-1基因的cDNA为模板,用Pfu酶做PCR扩增,用EcoR Ⅰ和HindⅢ双酶切,将目的基因克隆到pUCm-T质粒测序.进一步克隆到pET-Trx蛋白表达载体质粒,优化蛋白表达条件.结果含有IFITMP-1基因的PCR产物约1000 bp.重组pUCm-T质粒经EcoRⅠ、HindⅢ双酶切,有相应大小的cDNA片段插入,测序结果显示其序列正确,证实目的基因的扩增、克隆成功,并成功克隆进pET-Trx蛋白表达载体,经IPTG诱导,在BL21（DE3）plysS中有融合蛋白表达.结论成功扩增克隆IFITMP-1基因,并成功表达该基因编码的蛋白,为进一步研究IFITMP-1基因在大肠癌中的作用奠定了基础.

PCR amplification, cloning and protein expression of interferon-inducible transmembrane protein-1 gene

OBJECTIVE: To study the PCR amplification, cloning and protein expression of interferon-inducible transmembrane protein-1 (IFITMP-1) gene. METHODS: With the cDNA fragment containing IFITMP-1 gene as template, IFITMP-1 gene was amplified using Pfu enzyme by means of PCR. After EcoRI and HindIII digestion, the target gene fragment was linked to pUCm-T plasmid and sequenced. The IFITMP-1 gene was cloned into pET-Trx protein expression plasmid, and the condition for protein expression was optimized. RESULTS: The length of the PCR product of IFITMP-1 gene-containing cDNA fragment was about 1000 bp. The IFITMP-1 gene was successfully inserted into pUCm-T plasmid with correct sequence and cloning of the IFITMP-1 gene into the pET-Trx protein expression plasmid was achieved. Expression of the fusion protein of pUCm-T plasmid and IFITMP-1 gene was detected after IPTG induction. CONCLUSION: Successful amplification and cloning of the IFITMP-1 gene and its protein expression may facilitate further study of the role of IFITMP-1 gene in colorectal cancer.