The SMRI RNA samples were treated with DNase I using RNase-Free DNase Set (QIAGEN K.K., Tokyo, Japan) at room temperature (20–25°C) for 10 min, and then clean-up was performed using RNeasy® MinElute® Cleanup Kit (QIAGEN). First-strand cDNA for use in real-time qPCR was synthesized with the SuperScriptIII First-Strand synthesis system for qRT-PCR (Life Technologies Japan Ltd.) with 100 ng purified total RNA according to the manufacturer's protocol.
