真核绿色荧光蛋白表达载体pEGFP-C3-MAGE-12的构建与表达  

Construction and Expression of eukaryotic green fluorescent protein expression vector pEGFP-C3-MAGE-12

作　　者：张国俊[1] 赵国强[2] 王华启[1] 张世杰[2] 王蕾[2] 蒋莉[1] 

机构地区：[1]郑州大学第一附属医院呼吸内科,河南郑州450052 [2]郑州大学基础医学院微生物与免疫教研室,河南郑州450052

出　　处：《中国实用内科杂志：临床前沿版》2006年第3期425-427,共3页

基　　金：河南省医学科技创新人才工程(2005-38)

摘　　要：目的构建黑色素瘤抗原基因-12(MAGE-12)绿色荧光蛋白表达载体pEGFP-C3-MAGE-12,并在真核细胞中表达。方法于2005-10～2005-12对郑州大学第一附属医院应用RT-PCR方法,从人肺癌组织中扩增出MAGE-12cDNA基因片段,经过酶切鉴定后,克隆至质粒载体(pGEM-Teasy),测序证实碱基序列无误后,再克隆至真核绿色荧光蛋白表达载体(pEGFP-C3)上,并转染真核细胞,观察其在真核细胞中表达。结果经酶切及基因序列分析验证,PCR扩增出944bp的MAGE-12基因并成功构建了真核绿色荧光蛋白表达载体pEGFP-C3-MAGE-12,该重组载体能够在真核细胞中广泛表达。结论成功构建真核绿色荧光蛋白表达载体pEGFP-C3-MAGE-12,为建立肿瘤细胞疫苗打下基础。Objective To construct eukaryotic tluorescent expression vector pEGFP - C3 - MAGE - 12 and induce the vector express in eukaryotic cells. Methods From Oct. 2005 to Dec. 2005, mRNA was extracted from human lung cancer pecimen, MAGE -12 gene fragment was amplified by reverse transcriptase polymerase chain reaction （RTPCR） . After the amplified product was examined by electrophoresis and sequence determination, this fragment was inserted into pEGFP - C3 fluorescent expression vector . Then the expression vector was transfected into eukaryotie cell. Results The restriction enzyme digestion and sequence analysis showed that recombined cloning vector pGEM -TMAGE-12 and expression vector pEGFP- C3 - MAGE - 12 had been constructed successfully, which can express in eukaryotic cell stably. Conclusion Constructing the expression vector pEGFP - C3 - MAGE - 12 successfully, which make the foundation for producing a new tumor cell vaccine.

关 键 词：黑色素瘤抗原-12 真核表达 绿色荧光蛋白 克隆 

分 类 号：R5[医药卫生—内科学]