Figure 5.
Performance of normalization methods in detecting the effect of mutations on enhancer activity. (a) Design of reporter construct. Cebpa(0) contains the luc2 gene driven by the proximal Cebpa promoter. Cebpa(7) contains an enhancer in addition to the proximal promoter [4]. Binding sites for C/EBP family transcription factors and Gfi1 are shown in the magnified view. C/EBP sites have been mutated in Cebpa(7m1). (b) The relative activity of Cebpa(0), Cebpa(7), and Cebpa(7m1) inferred by either the REIV (left panels) or the ratiometric (right panels) methods in uninduced IL3 (progenitor) or induced GCSF (neutrophil) conditions. The activity has been normalized against the activity of the proximal promoter (Cebpa(0)). Error bars are standard errors of the mean. The ratiometric method spuriously detects an activation of Cebpa(7m1) in GCSF conditions (bottom right panel; Welch two sample t-test, , ).