siRNA specific to Sp1 reduces the transcriptional activity of the Dlk2 promoter and the expression of its mRNA. A) NIH3T3 cells were transiently transfected with the luciferase construct pGL3Basic(-212/+427), along with either pCMVSport (control) or pCMVSport-Sp1, to drive the overexpression of Sp1, and with a Sp1 siRNA or its negative control. Luciferase activities were measured 48 hours after transfection. In each transfection, luciferase values were normalized to the corresponding Renilla values, as previously described. B) NIH3T3 cells were transiently transfected with 10 nM of a Sp1-specific siRNA or with the corresponding negative control (Life Technologies). 48 hours after transfection the Sp1 and Dlk2 mRNA expression levels were measured by RT-qPCR. mRNA levels were referred to the expression level of phosphoriboprotein P0, used as an internal control, and the normalized values were referred to the values obtained with the negative control. The average values of three independent experiments are shown. (*, ** and ***, significant versus control in Student's t-test with p-values < 0.05, < 0.01 and < 0.001, respectively).