干扰素诱导的跨膜蛋白-1基因的扩增、克隆及蛋白表达

南方医科大学学报 ›› 2005, Vol. 25 ›› Issue (10): 1221-1224.

干扰素诱导的跨膜蛋白-1基因的扩增、克隆及蛋白表达

刘宇虎¹, 钟东², 柳娟¹, 张振书², 陈村龙², 武金宝², 肖冰², 郭文英¹

1. 东莞市人民医院消化科, 广东, 东莞, 523018;
2. 南方医科大学南方医院消化病研究所, 广东, 广州, 510515

出版日期:2005-10-20 发布日期:2005-10-20
基金资助:
收稿日期:2005-6-15。
基金项目:国家自然科学基金(30171053);广东省医学科学技术研究基金(B2005089);东莞市科技计划项目(B200501)
作者简介:刘宇虎(1963- ),男,2003年毕业于第一军医大学,博士,副主任医师,电话:0769-2223412,E-mail:liuyuhu@126.com

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

LIU Yu-hu¹, ZHONG Dong², LIU Juan¹, ZHANG Zhen-shu², CHEN Cun-long², WU Jin-Bao², XIAO Bing², GUO Wen-ying¹

1. 东莞市人民医院消化科, 广东, 东莞, 523018;
2. 南方医科大学南方医院消化病研究所, 广东, 广州, 510515

Online:2005-10-20 Published:2005-10-20

摘要/Abstract

摘要： 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EcoRⅠ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1000bp。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。

Abstract: 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 EcoRⅠand 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.

中图分类号:

刘宇虎¹, 钟东², 柳娟¹, 张振书², 陈村龙², 武金宝², 肖冰², 郭文英¹. 干扰素诱导的跨膜蛋白-1基因的扩增、克隆及蛋白表达[J]. 南方医科大学学报, 2005, 25(10): 1221-1224.

LIU Yu-hu¹, ZHONG Dong², LIU Juan¹, ZHANG Zhen-shu², CHEN Cun-long², WU Jin-Bao², XIAO Bing², GUO Wen-ying¹. PCR amplification, cloning and protein expression of interferon-inducible transmembrane protein-1 gene[J]. Journal of Southern Medical University, 2005, 25(10): 1221-1224.

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