%0 Journal Article %A Esmaeili, Fariba %T Enhancement of RNA Interference Effect in P19 EC Cells by an RNA-dependent RNA Polymerase %J Iranian Biomedical Journal %V 13 %N 1 %U http://ibj.pasteur.ac.ir/article-1-51-en.html %R - %D 2009 %K Gene silencing, RNA interference (RNAi), RNA-dependent RNA polymerase (RdRP), Double-stranded RNA (dsRNA), %X Background: RNA interference (RNAi) is a phenomenon uses double-stranded RNA (dsRNA) to specifically inhibit gene expression. The non-specific silencing caused by interferon response to dsRNA in mammalian cells limits the potential of utilizing RNAi to study gene function. Duplexes of 21-nucleotide short interfering dsRNA (siRNA) inhibit gene expression by RNAi. In some organisms, siRNA can also function as a primer converting mRNA into dsRNA that are further cleaved to produce more siRNA. This activity involves the enzyme RNA-dependent RNA polymerase (RdRP). There are no known RdRP involved in RNAi in mammals. By using an RdRP from Caenorhabditis elegance named ego-1, investigators intend to enhance RNAi effect in mammalian cells. The aims of this project were: 1) to investigate the efficiency of siRNA to enhanced green fluorescent protein (eGFP) gene silencing and 2) to enhance the RNAi effect. Methods: We used a vector-based siRNA to target eGFP. Also we used a vector expressing ego-1 to test for a possible amplification effect of RNAi. The expression of eGFP in the cells was detected by using fluorescent microscopy, flowcytometry and Western-blotting. Results: Transfection of the plasmid into P19 cells significantly decreased eGFP fluorescence. In addition, eGFP protein was reduced. Preliminary data suggested that the presence of ego-1 enhanced the RNAi effect. Conclusion: The results indicated that use of hairpin siRNA expression vectors for RNAi is a promising method to inhibition of gene expression in mammalian cells. Also, introducing RdRP enzyme to mammalian cells might amplify the RNAi effect in the cells. %> http://ibj.pasteur.ac.ir/article-1-51-en.pdf %P 19-25 %& 19 %! Enhancement of RNA Interference %9 Full Length/Original Article %L A-10-1-16 %+ %G eng %@ 1028-852X %[ 2009