Real-Time Analysis of mRNA Expression

Total mRNA was extracted with TRIzol (Invitrogen, Carlsbad, CA, USA). mRNA was reverse transcribed into cDNA using a Transcript or First-Strand cDNA Synthesis Kit (Roche Life Science, Basel, Switzerland) according to the manufacturer's directions. The SYBR Green real-time PCR method was used to compare gene expression. The primers are shown in Table 2. Changes in mRNA expression were normalized to that of actin and were calculated using the 2^−ΔΔCT method as previously described.¹⁶

Primers for RT-PCR