Inhibition of EGFP expression by siRNA in EGFP-stably expressing Huh-7 cells.
The RNA interference (RNAi) is a powerful tool to silence gene expression post-transcriptionally, and causes the degradation of an mRNA containing the same sequence. In this present study, an alternative approach was used to in vitro synthesize enhanced green fluorescent protein (EGFP) specific short interfering RNA (siRNA) using T7 RNA polymerase, and a pEGFP-N1 transfected, human hepatoma cell line Huh-7 derived Huh-7-N cell clone was established. When introduced the siRNA into the EGFP expressing Huh-7-N cells, the EGFP specific siRNA was able to specifically inhibit the expression of EGFP in Huh-7-N. In comparison with that in wild-type Huh-7 or that in Huh-7 co-transfected with pEGFP-N1, the inhibition of EGFP specific siRNA in Huh-7-N cells is more significant and repeatable. It is concluded that a cell clone Huh-7-N, which stably expresses EGFP, has been established, and the in vitro synthesized EGFP siRNA can be used in silencing the EGFP gene expression. This Huh-7-N/EGFP specific siRNA system has been proved reliable and convenient, and can also be applied widely as control in other RNA interference studies.
Cao
Mingmei
M
Department of Microbiology, Second Military Medical University, Shanghai 200433, China.
Ren
Hao
H
Pan
Xin
X
Pan
Wei
W
Qi
Zhong-Tian
ZT
D002460:Cell Line; D015870:Gene Expression / Q000187:drug effects*; D020868:Gene Silencing / Q000187:drug effects*; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology*; D014162:Transfection
D008164:Luminescent Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D049452:Green Fluorescent Proteins