We are interested in identifying compounds that can either increase or decrease the expression of our gene of interest. We usually seed cells in 384 well plates and then incubate cells with stress inducer in presence and absence of compounds. Due to enormous number of compounds to be screened I would like to isolate RNA directly from 384 well plate for processing.
Can you follow gene expression using a reporter cell assay, in which the promoter of the gene of interest is placed upstream of a reporter such as GFP or luciferase? Then you would just measure the effect of the compound on the fluorescence or luminescence of the cells, instead of extracting the RNA.
Thanks Dr. Shapiro..I have been using reporter assays to narrow down the list but I still think it will be a bit time consuming to develop reporter cell assay for each gene rather than isolating RNA and performing assay for multiple genes at the same time.
One idea is to break open the cells to release the mRNA, then capture polyadenylated mRNA with biotinylated oligo dT that is attached to streptavidin-coated beads. The RNA could then be released into solution from the beads by adding polydA or free biotin.
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