Figure 5.

Relative promoter inducibility is maintained across plasmid copy numbers and upon genomic integration. (a) HEK 293T cells were transiently transfected with various concentrations of the sfGFP-expressing plasmids shown in Figure 2a. Fold-induction was calculated based on median sfGFP intensity among dsRed+ gated cells. (b–c) HEK 293T cells were lentivirally transduced with constructs encoding sfGFP expressed from various hypoxia-inducible promoters. Transduced cells were identified by the expression of EGFRt, which was expressed from a constitutive EF1α promoter encoded in the same lentiviral vector. (b) Median sfGFP intensity of EGFRt+ gated cells was quantified by flow cytometry. (c) Fold-induction was calculated from the data shown in (b). The large error bar for fold-induction of YB_TATA resulted from the fact that this core promoter had very low basal expression (i.e., small denominator in the fold-induction calculation; see Figure 5b). Slight variations in basal gene expression resulted in a large standard deviation in fold-induction after error propagation. Values shown are the means of triplicates with error bars indicating ± 1 s.d.