摘要
目的应用载体介导的RNA干扰(RNA i)技术特异性地抑制甲状腺癌细胞人端粒酶逆转录酶(hTERT)基因的表达。方法构建利用U6启动子转录功能性小干扰RNA(siRNA)的质粒载体,对阳性克隆进行酶切和测序鉴定后,转染人甲状腺癌FRO细胞,利用RT-PCR和W eston blot法检测hTERT基因的表达情况。结果构建的表达质粒转染FRO细胞后,hTERT基因mRNA和蛋白质的表达水平明显降低,抑制率分别为76%和81%。结论siRNA的表达质粒能成功抑制甲状腺癌细胞hTERT基因的表达,为RNA i技术应用于甲状腺癌的基因治疗提供了实验基础。
Objective To inhibit the expression of human telomerase reverse transcriptase gene （hTRET） in thyroid cancer cells by using vector - based RNA interference （ RNAi ） technique. Methods A vector containing U6 promoter used to transcribe functional small interfering RNA （siRNA） targeting hTRET gene was constructed, After identification by restriction enzyme digestion and sequencing, the plasmid of positive clones was transfected into human thyroid carcinoma FRO cell line, The expression of hTRET gene was detected by RT - PCR and Western blot. Results After the recombinant siRNA expression plasmid transfected into FRO cells, the mRNA and protein expression of hTRET gene were significantly suppressed and the inhibition ratio reached 76% and 81% respectively. Conclusion The siRNA expression plasmid targeting hTRET gene could specially suppress its expression in thyroid carcinoma cell. This provides a experimental basis for RNA interference technique in gene therapy on thyroid carcinoma.
出处
《中国全科医学》
CAS
CSCD
2006年第4期290-292,共3页
Chinese General Practice
关键词
RNA干扰
甲状腺肿瘤
人端粒酶逆转录酶
质粒
RNA interference
Thyroid neoplasms
Human telomerase reverse transcriptase
Plasmid
