RT-PCR Gene Expression Validation

RT-PCR was performed in order to obtain cDNA using the Moloney murine leukemia virus-reverse transcriptase (MMLV-RT) kit (Life Technologies. Marseille. France) as previously described [18 (link)]. Specific genes found modulated by microarray or strongly involved in lymphoma according to prior work² were then selected and validated by q-PCR using specific primers listed in the S1 Table as well as the SYBR Green Fast Master Mix (Roche Diagnostics, Meylan, France). The results were normalized to housekeeping gene β-actin and expressed as fold change (FC) = 2^−ΔΔCt, which ΔΔCt = (Ct_Target—Ct_Actin)_assay—(Ct_Target—Ct_Actin)_control. The threshold cycle (Ct) was defined as the number of cycles required to detect the fluorescent signal. The expression of genes was considered as modulated when FC ≥ 1.5.