E-GEOD-6697 - Transcription profiling of primary mouse embryonic fibroblasts (MEF) in response to serum
Submitted on 10 January 2007, released on 15 June 2008, last updated on 27 March 2012
Current methods to analyze gene expression measure steady-state levels of mRNA. In order to specifically analyze mRNA transcription, a technique has been developed that can be applied in-vivo. The technique is referred with the acronym NIAC-NTR (Non Invasive Application and Capture of Newly Transcribed RNA). This method makes use of the cellular pyrimidine salvage pathway and is based on affinity-chromatographic isolation of thiolated mRNA. When combined with data on mRNA steady-state levels, this method is able to assess the relative contributions of mRNA synthesis and degradation/stabilization. It overcomes limitations associated with currently available methods such as mechanistic intervention that disrupts cellular physiology, or the inability to apply the techniques in-vivo. The method has been applied to a model of serum response of cultured primary mouse embryonic fibroblasts. Affymetrix GeneChip microarrays were used to detail regulatory mechanisms of mRNA expression and the relative contributions of RNA synthesis and turnover within distinct pathways, and identification of genes expressed at low abundance at the transcriptional level. Experiment Overall Design: The gene expression and regulation program at the RNA level early (2h) in response to serum was studied. Total RNA, amplified total RNA and specifically enriched newly transcribed RNA was isolated from serum starved and 2h serum treated MEF cells. This experiment allowed detailed regulatory mechanisms of mRNA expression and the relative contributions of RNA synthesis and turnover within distinct pathways, and identification of genes expressed at low abundance at the transcriptional level of MEF cells early in response to serum.
transcription profiling by array, unknown experiment type
Microarray analysis of newly synthesized RNA in cells and animals. M Kenzelmann, S Maertens, M Hergenhahn, S Kueffer, A Hotz-Wagenblatt, L Li, S Wang, C Ittrich, T Lemberger, R Arribas, S Jonnakuty, M C Hollstein, W Schmid, N Gretz, H J Gröne, G Schütz. Proc Natl Acad Sci U S A 104(15):6164-9 (2007)