AIF重组质粒的构建、表达和功能的研究  

Preliminary studies on construction,expression and function of recombinant AIF plasmid

作　　者：袁长青[1] 丁振华[1] 马树东[1] 凌朝辉 刘胜军[1] 胡贵方[1] 郑莉[1] 周美娟[1] 欧程山[1] 

机构地区：[1]第一军医大学热带军队卫生学系放射医学教研室,广东广州510515 [2]上海转基因研究中心

出　　处：《毒理学杂志》2005年第3期202-204,共3页Journal of Toxicology

基　　金：国家自然科学基金资助(30200226)

摘　　要：目的构建人凋亡诱导因子(AIF)基因功能片段的真核表达载体pcDNA3.1(+)AIF,并初步研究AIF在细胞凋亡中的作用。方法从人细胞中提取总RNA,RTPCR扩增AIF基因片段;克隆到pMD18T载体上,通过酶切和测序进行鉴定;将AIF基因片段插入pcDNA3.1(+),构建真核表达载体;电转化淋巴母细胞Raji细胞,观察AIF的表达水平和功能。结果成熟AIF蛋白分子在真核细胞中表达,并转移到细胞核中,诱导Caspase非依赖性细胞凋亡。结论成功构建了pcDNA3.1(+)AIF真核表达载体,AIF在Caspase非依赖性细胞凋亡中发挥重要作用。Objective To construct eukaryotic expression vector carrying human Apoptosis-inducing factor（AIF） gene fragment, pcDNA3.1（ ＋ ）/AIF,and research the gene expression and function. Methods After total RNA was extracted from human cells,AIF gene fragment was amplified by RT-PCT,then cloned into pMD18-T vector,which was certified by restriction analysis and sequencing. The gene fragment was inserted into plasmid pcDNA3.1 （ ＋ ） and eukaryotic expression vector was constructed. The vector was transfected into Raji cells, and then the gene expression and function was studied. Results Eukaryotic expression vector, pcDNA3.1 （ ＋ ）/AIF,was constructed and the gene was expressed in eukaryotic cells successfully. The expressed AIF protein was translocated into nucleus and induced caspase independent apoptosis. Conclusion AIF plays an important part in caspase-independent apoptosis.

关 键 词：凋亡诱导因子 真核表达载体 构建 AIF PCDNA3.1(+) Caspase非依赖性 重组质粒 RT-PCR扩增 细胞凋亡 基因片段 

分 类 号：R735.7[医药卫生—肿瘤] R644[医药卫生—临床医学]