Figure 4

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).