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作 者 ：潘晓菲, 时丽丽, 谭初兵, 吕超君, 徐为人, 汤立达

期 刊 ：生物技术通报 2014年 0卷 2期 页码：187-192

关键词：内皮素受体A；RT-PCR；pTag-Lite；SNAP载体；CHO-K1细胞；表达；

摘 要 ：为了建立内皮素受体拮抗剂筛选系统，克隆人ETA基因，构建真核表达载体，并实现pTag-Lite SNAP-ETA在CHO-K1细胞中的瞬时表达。从人肺腺癌细胞系A549中克隆人ETA基因，连接到pTag-Lite SNAP质粒，构建表达载体pTag-Lite SNAP-ETA，用FuGENER HD转染试剂将表达载体pTag-Lite SNAP-ETA转染入CHO-K1细胞内，通过荧光显微镜检测融合蛋白SNAP-ETA的表达。DNA测序结果表明pTag-Lite SNAP-ETA表达载体构建成功，荧光显微镜检测结果表明人ETA在CHO-K1细胞中有效表达。

Abstract:It was to facilitate inhibitor screening of endothelin receptor, cloning of human endothelin receptor gene ETA, construction of eukaryotic expression vectors and transient expression of pTag-Lite SNAP-ETA in CHO-K1 cells. Method Human ETA gene fragment were amplified from human lung cancer cell line A549 via RT-PCR and inserted into pTag-Lite SNAP plasmid, the recombinant plasmids were then transiently transfected into CHO-K1 cells using FuGENE? HD, expression of ETA was observed via fluorescence microscope. Result Sequencing result showed pTag-Lite SNAP-ETA expression vector were correctly constructed. Fluorescence microscope images indicated that two vectors were expressed in CHO-K1 cells.

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