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- W1503784758 abstract "Top of pageAbstractSilencing of expression produced by small interfering RNA (siRNA) directed against different regions of a lentiviral vector transgene expressing enhanced green fluorescent protein (EGFP) driven by the elongation factor 1 alpha (EF1a) promoter 12 was investigated in transfected and transduced 293FT fibroblasts and human peripheral mononuclear cells. SiRNA directed against exonic sequences significantly reduced expression, including one targeting exon 1 (+10 to +30 relative to the transcriptional start site), and one targeting EGFP coding sequences (+1167 to +1185). Unexpectedly, siRNA directed against intron A (+155 to +175 relative to the transcriptional start site) or targeting the EF1a promoter (−106 to −86 and −25 to −5 start) also silenced EGFP expression. Post-transcriptional gene silencing in human cells has been described to be restricted to the cytoplasm 3, and transcriptional gene silencing (TGS) mediated by RNA interference (RNAi) has not previously been shown to occur in mammalian cells. Silencing of expression could not be attributed to reduced vector transduction, non-specific inhibition of transcription, cytotoxicity, initiation from a cryptic promoter, or aberrant splicing of EGFP mRNA, and was confirmed to occur at the level of transcription by nuclear run-on. Endogenous EF1a expression could inhibited by transfecting siRNA using a reagent (MPG)4w which facilitates nuclear entry. Inhibition of expression and methylation of one site in the promoter was reversible with addition of 5′ azacytidine and trichostatin A. Treatment with siRNA targeting intronic sequences and promoter regions can induce transcriptional gene silencing. Access of siRNA to the nucleus may be key to the observed effects." @default.
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- W1503784758 date "2004-05-01" @default.
- W1503784758 modified "2023-10-06" @default.
- W1503784758 title "RNA mediated transcriptional gene silencing in human cells" @default.
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- W1503784758 doi "https://doi.org/10.1016/j.ymthe.2004.06.240" @default.
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