摘要
为了分析在美洲棉铃虫细胞(HzAM1)内RNAi的效果,将egfp基因克隆到含有双向T7启动子/终止子的质粒载体中,在体外合成全长的增强型绿色荧光蛋白(egfp)基因dsRNA,将dsRNA和含有能在昆虫细胞内表达eGFP的质粒一起转染HzAM1细胞,分析dsRNA对eGFP表达的抑制作用。结果显示,由egfp基因转录的长dsRNA能有效抑制HzAM1细胞内eGFP的表达,而且该抑制作用表现为剂量依赖效应。但是抑制作用并不彻底,在高剂量的dsRNA处理下,仍有部分细胞内能观察到eGFP的表达。
Double-stranded RNA-mediated interference（RNAi） has recently emerged as a powerful reverse genetics tool to silence gene expression in multiple organisms,including plants,nematodes and insects.In this study,DNA vector capable of promoting the synthesis of full-length dsRNA of egfp gene in vitro have been constructed,in which egfp gene was inserted into the bidirectional T7 promoter-terminator region.The synthesized dsRNA and a plasmid containing egfp cassette were transfected into HzAM1 cells to analyze eGFP expression silencing.Results showed that the synthesized dsRNA was able to inhibit eGFP expression in a dose dependent manner.However,the gene expression could not be completely knocked down.There still were a few cells were detected eGFP expression post-treatment with high dose dsRNA.Those results provide a reference for RNAi in insect cells.
出处
《生物技术通报》
CAS
CSCD
北大核心
2011年第11期130-133,共4页
Biotechnology Bulletin
基金
国家自然科学基金项目(30970138)
