Inhibition of EGFP expression by siRNA in EGFP-stably expressing Huh-7 cells.

Abstract

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.

• Authors: Mingmei Cao, Hao Ren, Xin Pan, Wei Pan, Zhong-Tian Qi
• Journal: Journal of virological methods (J Virol Methods) Vol. 119 Issue 2 Pg. 189-94 (Aug 2004) ISSN: 0166-0934 [Print] Netherlands
• PMID: 15158602 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Luminescent Proteins
• RNA, Messenger
• RNA, Small Interfering
• Green Fluorescent Proteins

Topics

• Cell Line
• Gene Expression (drug effects)
• Gene Silencing (drug effects)
• Green Fluorescent Proteins
• Humans
• Luminescent Proteins (genetics, metabolism)
• RNA, Messenger (drug effects, metabolism)
• RNA, Small Interfering (pharmacology)
• Transfection