Lightcycler 2.0 2

Total RNA was extracted using the Ambion TRIzol Reagent (Thermo Fisher Scientific, Waltham, MA, USA), and complementary DNA was synthesized from 1.0 μg of the total RNA by using the oligo dT primer and a reverse transcription system (Cat. #A3500, Promega, Madison, WI, USA) according to the manufacturer’s instructions. A quantitative real-time polymerase chain reaction (qRT-PCR) was performed using Light Cycler Faststart DNA Master SYBR Green I (Roche Diagnostics, Indianapolis, IN, USA) and a LightCycler TaqMan Master (Roche Diagnostics, Indianapolis, IN, USA) on LightCycler 2.0 II (Roche Molecular Systems, Branchburg, NJ, USA). Intergroup differences in target gene expression were determined using the 2^−ΔΔCt method.

Total RNA was collected from cultured cells or tumor tissues using TRIzol Reagent (Thermo Fisher Scientific), and complementary DNA was synthesized from 1.0 μg of total RNA using oligo dT primer and a Reverse Transcription System (Promega, Madison, WI, USA) according to the manufacturer’s instructions. Real-time RT-PCR was carried out using LightCycler FastStart DNA Master SYBR Green I (Roche, Basel, Switzerland) on a LightCycler 2.0 II (Roche). Expression of the target gene was normalized relative to GAPDH mRNA expression using the 2^−ΔΔCt method. Primers are shown in Supplementary Table S1.