RT-qPCR validation of EGF-regulated miRNAs. Total RNA prepared from cells lysates were analyzed by quantitative real time PCR using the miRCURY LNA™ microRNA PCR System (Exiqon) for each of the miRNAs as indicated. A. Control treatment without inhibitors: HeLa cells were serum-starved for 24 hours and treated with EGF for 1 and 6 hours. B. Inhibitor treatment. HeLa cells were serum-starved for 24 hours and treated with EGF for 6 hours in the presence or absence of protein kinase inhibitors: AG1470 (EGFR inhibitor), U0126 (MEK inhibitor) and Wortmannin (PI3K inhibitor). In addition, HeLa cells were transfected with a constitutively active form of Ras (RasV12). Effective pathway inhibition was verified by western blotting in parallel samples from the same experiment (see Additional file 8).
Llorens et al. BMC Genomics 2013 14:371 doi:10.1186/1471-2164-14-371