水稻重复序列RRD3在CHO和3T3细胞中的启动子活性

RRD3是通过变性 -复性动力学从水稻基因组中克隆到的一个中度重复序列 .将 RRD3作为启动子亚克隆到氯霉素乙酰基转移酶基因为报告基因的 p CAT载体上 ,转染 5种动物细胞 .结果表明在 CHO、3T3细胞中能够启动 CAT基因的表达 .利用核酸外切酶 和 PCR构建了 RRD3的系列缺失体 ,经转染后的 CAT活性分析表明 ,该序列中存在多个与哺乳动物细胞中启动基因表达有关的调控元件 .结合 RRD3在大肠杆菌和高等植物烟草中启动报告基因表达的活性 ,对重复序列与生物进化的关系进行了讨论 .

The Promoter Activity of RRD3 in CHO and 3T3 Cell Lines *

The RRD3 repetitive sequence was cloned from the rice genome according to the characteristics of repetitive sequences during denature anneal procedure.This repetititive sequence showed great tetracycline resistance in E.coli .RRD3 was subcloned into the mammalian report vectors.It displayed different promoter activity in the mammalian cell lines,CHO and 3T3,according to the respective CAT activity that had been detected.In order to locate the exact promoter region in this sequence,6 sequence deletants were constructed with exonuclease Ⅲ and PCR.The CAT assay after the transfection of deletants demonstrated that there were several DNA functional subsequences within this repetitive sequence.Furthermore,the relation between repetitive sequences and species evolution was also discussed here with respect to the promoter activity of RRD3 in E.coli ,plant (tobacco and rice) and mammalian cell lines.