【摘 要】
：
目的:采用flag标记的人FAT10蛋白研究人FAT10蛋白对HEK293细胞的影响.方法:将FAT10基因片段克隆到载体pcDNA3-flag上,对阳性克隆进行PCR、酶切和测序鉴定,用脂质体转染HEK293
【机 构】
：
南方医科大学广东省蛋白质组学重点实验室和病理生理学教研室
论文部分内容阅读
目的:采用flag标记的人FAT10蛋白研究人FAT10蛋白对HEK293细胞的影响.方法:将FAT10基因片段克隆到载体pcDNA3-flag上,对阳性克隆进行PCR、酶切和测序鉴定,用脂质体转染HEK293细胞,用Western blotting方法检测外源性FAT10正常培养状态和饥饿状态下的HEK293细胞中的表达情况;并用XTT法和DNA ladder法观察正常培养和饥饿状态下HEK293细胞的凋亡情况.结果:重组质粒在HEK293细胞中高效表达,但在正常培养状态下和饥饿状态下表达情况不同.饥饿状态下,过表达FAT10的HEK293细胞存活率显著低于对照细胞,并且出现DNA ladder现象.结论:成功构建了带Flag标签的FAT10真核表达质粒,可在HEK293细胞中高效表达;FAT10过表达促进饥饿状态的HEK293细胞发生凋亡.
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