Sybr green 1 system

RNA was isolated using Trizol (Invitrogen) and reverse transcribed with Superscript II (Invitrogen) per manufacturer protocols. Quantitative real-time polymerase chain reaction analysis of target gene expression was performed using the iCycler and SYBR Green I system (Bio-Rad) as described[12 (link)]. Samples were analyzed in triplicate and normalized to expression values of mouse housekeeping gene TFIIB or human housekeeping gene TBP. Data were calculated using the 2-ΔΔCT method[25 (link)]. The following primer sequences were used: mouse NOR1 (forward: 5′-AGACGCCGAAACCGATGT-3′ and reverse: 5′-TCGGACAAGGGCATTCA-3′), mouse RUNX1 (forward: 5′-GCAGGCAACGATGAAAACTACT-3′ and reverse: 5′-GCAACTTGTGGCGGATTTGTA-3′), mouse TFIIB (forward: 5′-CTCTCCCAAGAGTCACATGTCC-3′ and reverse: 5′-CAATAACTCGGTCCCCTACAAC-3′), human RUNX1 (forward: 5′-TCTTCACAAACCCACCGCAA-3′ and reverse: 5′- CTGCCGATGTCTTCGAGGTTC-3′), human NOR1 (forward: 5′-GGGCTTTTTCAAGAGAACAGTG-3′ and reverse: 5′-ATCTCTGGGTGTTGAGTCTGTT-3′), human TBP (forward: 5′-GGAGAGTTCTGGGATTGTACCGC-3′ and reverse: 5′-ATATTCGGCGTTTCGGGCAC-3′).