Quantitative PCR Analysis of Transcription Factors

Quantitative PCR was performed using the Rotor-Gene SYBR Green PCR kit (Qiagen, Hilden, Germany), The primer sequences used include RelB: forward, 5′-GTTCCAGTGACCTCTCTTCC-3′, reverse, 5′-CCAAAGCCGTTCTCCTTAATGTA-3′,^{18 (link)}Maf forward, reverse,^{18 (link)}GM-CSFRα: forward, 5′-CTGATGTCATGAAGCGATGC-3′, reverse, 5′-TCACGGTGACATCAATGTCG-3′,^{19 (link)}PU.1: forward, 5′-GATGGAGAAGCTGATGGCTTGG-3′, reverse, 5′-TTCTTCACCTCGCCTGTCTTGC-3′,^{20 (link)}C/EBPα: forward, 5′-TGGACAAGAACAGCAACGAGTAC-3′, reverse, 5′-GCAGTTGCCCATGGCCTTGAC-3′^{21 (link)} and GAPDH: forward, 5′-TCCACCACCCTGTTGCTGTA-3′, reverse, 5′-AATGTGTCCGTCGTGGATCT-3′. The rotor-gene Q (2plex on PC) instrument (Qiagen) was programmed as follows: 95 °C for 10 min, then 50 cycles of 95 °C for 10 s, 60 °C for 15 s and 72 °C for 20 s. The data were analyzed using rotor-gene Q series software through the delta–delta CT method.^{22 (link)}