Rt2 profiler pcr data analysis software version 3

RNA isolation from cultured cells was performed using RNeasy® mini kit (Qiagen, USA) according to the manufacturer’s protocol. RNA Samples were stored at −80°C until further use. For cDNA preparation, 1μg of total RNA was reverse transcribed at 37°C using high capacity cDNA kit according to manufacturer`s instructions (Applied Biosystems, Foster City, CA). For cell death pathway analysis (RT2 profiler, SA Biosciences, Frederick, MD, USA), PCR array based expression profiling was performed using SYBR-Green method and the results were analyzed using the ΔΔCt method using RT² profiler PCR data Analysis software version 3.5 (SA Biosciences).

The OVCA cells were washed with PBS and detached using accutase solution (Sigma–Aldrich, St Louis, MO, USA). For RNA extraction, RNeasy® mini kit (Qiagen, Valencia, CA) was used according to the manufacturer's instruction. Reverse transcription was performed using the high capacity cDNA kit (Applied Biosystems, Foster City, CA) according to manufacturer's protocol. All real-time PCR reactions were performed in triplicate in 10 μl volume using Universal fast PCR Master Mix reagent (Applied Biosystems, USA) according to the manufacturer's instructions. The results were analyzed using the ΔΔCt method using GAPDH as the endogenous control. For cell death and autophagy pathway analysis (RT2 profiler, SA Biosciences, Frederick, MD, USA), PCR array-based expression profiling was performed using SYBR-Green method and the results were analyzed using the ΔΔCt method using RT² profiler PCR data analysis software version 3.5 (SA Biosciences).