Stable suppression of gene expression by short interfering RNAs targeted to promoter in a mouse embryonal carcinoma stem cell line

Esmaeili, F. and Bamdad, T. and Ghasemi, S. (2010) Stable suppression of gene expression by short interfering RNAs targeted to promoter in a mouse embryonal carcinoma stem cell line. In Vitro Cellular and Developmental Biology - Animal, 46 (10). pp. 834-840.

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Abstract

RNA interference (RNAi) can induce gene silencing via two pathways: post-transcriptional gene silencing (PTGS) and transcriptional gene silencing (TGS). The mediators of gene inactivation in both pathways are 21-bp small interfering RNAs (siRNAs) generated from longer double-stranded RNA (dsRNA). PTGS involves siRNA-mediated targeting and degradation of mRNA. However, siRNAs induce TGS via DNA methylation at the targeted promoter. Synthetic siRNAs can induce loss of gene activity comparable to long dsRNA. The limitation of this method is that the transfected synthetic siRNA works for only a few days. In this study, we tested the RNAi response to siRNA (PTGS pathway) by using a plasmid containing an enhanced green fluorescent protein (eGFP) gene as a target as well as a plasmid creates siRNA transcript, in a form of a hairpin, against eGFP gene. To investigate TGS pathway via RNAi, we also used a plasmid creates hairpin siRNA transcript against pgk-1 promoter. The data presented here indicated long-lasting inhibition in expression of eGFP and puromycin genes, both under the control of the murine Pgk-1 promoter. However, Southern blot analysis showed no methylation in pgk-1 promoter. © 2010 The Society for In Vitro Biology.

Item Type: Article
Additional Information: cited By
Uncontrolled Keywords: enhanced green fluorescent protein; green fluorescent protein; small interfering RNA, animal; animal embryo; article; gene silencing; genetics; metabolism; mouse; plasmid; RNA interference; Southern blotting; tumor cell line, Animals; Blotting, Southern; Cell Line, Tumor; Embryo, Mammalian; Gene Silencing; Green Fluorescent Proteins; Mice; Plasmids; RNA Interference; RNA, Small Interfering, Murinae
Subjects: QZ pathology-Neoplasms
QU Biochemistry > Cell biology and genetics
Divisions: Reserach Vice-Chancellar Department > Cellular and Molecular Research Center
Depositing User: zahra bagheri .
Date Deposited: 20 Aug 2017 08:19
Last Modified: 20 Aug 2017 08:19
URI: http://eprints.skums.ac.ir/id/eprint/3311

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