期刊论文

中文

中国动脉硬化杂志

1007-3949

2012

20

5

397-401

国家自然科学基金项目（81000035、31071999和30871340）；

本校为第一机构

目的 利用siRNA表达载体构建靶向人类心脏发育候选基因hole的pSUPER RNAi载体系(pSUPERhole)及筛选稳定产毒的细胞克隆.方法 化学合成一对编码短发夹RNA序列的、靶向人类心脏发育候选基因hole的寡核苷酸链60个碱基,退火,克隆到经BglⅡ、Xho Ⅰ双酶切的pSUPER质粒上,构建重组RNAi质粒(pSUPER-hole).通过双酶切鉴定及测序分析验证构建效果.将正确构建的质粒转染大鼠心肌细胞H9c2,经过puro抗生素的筛选,建立稳定产生逆转录病毒的细胞模型,采用RT-PCR检测其抑制效果.结果 pSUPER-hole载体经双酶切鉴定及测序分析,结果表明60个碱基成功插入到预计位点,并且序列完全一致.重组载体转染包装细胞,筛选的细...

Aim To construct a pSUPER RNAi system that targets human heart developmental candidate gene hole and a stable virus-producing cell line. Methods A pair of 60nt oligonucleotides coding for short hairpin RNA and targeting human heart developmental candidate gene hole were chemically synthesized and annealed and inserted into pSUPER plasmids digested with Bgl II and Xho I to construct the recombinant pSUPER RNAi plasmid （pSUPER-hole）. Recombinant pSUPER-hole plasmid was identified by enzyme digestion and sequencing analysis. The packaging cell H...