VEGFR-3胞外区基因真核表达载体的构建与鉴定  

Construction and Identification of Eukaryotic Expression Vector of VEGFR-3 Extracellular Domain Gene

作　　者：陈艳[1] 王熙才[1] 伍治平[1] 金从国[1] 谷玉兰[1] 周永春[1] 刘馨[1] 

机构地区：[1]昆明医学院第三附属医院,云南省肿瘤医院/肿瘤研究所,650118

出　　处：《肿瘤防治研究》2009年第1期40-43,共4页Cancer Research on Prevention and Treatment

基　　金：云南省科技厅;昆明医学院联合专项基金资助项目(2007C0024R)

摘　　要：目的构建VEGFR-3胞外区基因真核表达载体pcDNA3.1-VR-3,并在体外进行表达和鉴定。方法采用RT-PCR技术,扩增C57BL/6小鼠胚胎VEGFR-3胞外区cDNA片段,通过基因重组技术将其插入到真核表达载体pcDNA3.1,构建重组质粒pcDNA3.1-VR-3。经限制性酶切鉴定和DNA序列测定结果证实后,将重组质粒经脂质体法转染COS-7细胞,Western blotting检测其蛋白表达。结果克隆了C57BL/6小鼠胚胎VEGFR-3胞外区cDNA片段,并构建了真核表达载体pcDNA3.1-VR-3,Western blot证实目的基因可在COS-7细胞中表达。结论构建的真核表达载体pcDNA3.1-VR-3,可在真核细胞内正确表达,这为进一步的动物实验奠定了基础。Objective To construct the pcDNA3. 1-VR-3 eukaryotic expression vector for VEGFR-3 extracellular domain gene. The expression and identification of the vector was carried out in vitro. Methods The extracellular domain of VEGFR-3 encoding sequence was amplified by reverse transcriptase-polymerase chain reaction from C57BL/6 mice embryo and cloned into the Hind Ⅲ-Xba Ⅰ sites of pcDNA3. 1. After confirmed by sequencing the recombinant plasmid pcDNA3. 1-VR-3 was transfected into COS-7 cells and its protein expression was identified by Western blot. Results The extracellular domains of VEGFR3 encoding sequence was successfully cloned from C57BL/6 mice embryo. And the pcDNA3. 1-VR-3 eukaryotic expression vector was constructed, which can be expressed in COS-7. Conclusion We successfully constructed the pcDNA3. 1-VR-3 eukaryotic expression vector which may pave a way for further studies in animals experiment.

关 键 词：VEGFR-3 淋巴生成 真核表达 

分 类 号：R73-75[医药卫生—肿瘤]