RNA isolation and quantitative real-time PCR

RNA isolation and qRT-PCR was performed as previously described (38). Briefly, total RNA was extracted using Trizol reagent according to the manufacturer's protocols (Invitrogen). 1 μg of RNA was used for reverse transcription using a high capacity DNA reverse transcription kit (Applied Biosystems Cat No: 4368813), and qRT-PCR was performed using Power SYBR green PCR master mix (Applied Biosystems #4367659) according to manufacturer's protocols. The fold-change in expression was calculated by the 2^–ΔΔCt methods. mRNA expression profiles were normalized to levels of housekeeping gene Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in each sample The primers used in qRT-PCR are listed in supplementary material and methods.