Knock down of target genes by induced expression of miR-122. a. Glycogen synthase-3' UTR targeted by miR-122. NIH3T3-47/X1-miR122 cells were transfected with reporter plasmids pTKGLuc (control), or pTKGLuc-GYS with the (glycogen synthase 3' untranslated region (UTR) and mIR-122 target sites schematic, top panel). The luciferase activity remaining 48 h after induction is plotted as a percent of activity from control cells. (*p = 0.0255; Error bar = -/+1SD) (See Methods for 3' UTR sequence coordinates.) b. Western blot analysis of aldolase A, an endogenous target of miR-122,. NIH3T3-47/X1-miR122 cells were treated with DMSO or 0.5 μM RSL1, then cell lysates were used for western blot analysis. The sample of 9 days post treatment is shown. Aldolase A protein quantification was calculated after LiCor scanning of Western blot normalized for loading with alpha-beta tubulin. A single miR-122 target site in aldo A is located at position 27-34 in the aldoA 3' UTR (top panel).
Shea and Tzertzinis BMC Biotechnology 2010 10:76 doi:10.1186/1472-6750-10-76