Skip to main content
Fig. 4 | Genome Biology

Fig. 4

From: A novel mechanism for variable phenotypic expressivity in Mendelian diseases uncovered by an AU-rich element (ARE)-creating mutation

Fig. 4

Effect of SCL4A4 ARE-forming mutation on mRNA decay. a Hap-1 fibroblast-like cells were transfected with nanoluciferase reporters fused with WT SLC4A4 3′UTR or the ARE-forming mutant -SLC4A4 3′UTR for 16 h. Total RNA was extracted and levels of reporter mRNA were quantitated by RT-QPCR using TaqMan primers specific to nanoluciferase and were normalized to the housekeeping gene, GAPDH. Data are mRNA ratio, mean ± SEM of triplicate measurements from two experiments. Statistical significance was assessed by Student’s t-test; ***P < 0.0001. b Hap-1 cells were transfected with either WT or mutant SLC4A4 3′UTR constructs. The cells were treated with actinomycin D (5 μg/mL) up to 6 h. RT-QPCR was performed on all samples and the relative abundance level of the reporter’s transcript was taken as a measure of the ratio between the construct and an endogenous gene (RPLPO). c HeLa Tet-off cells (3 × 104 cells/well) were transfected with TetO-linked reporters fused with WT SLC4A4 3′UTR or the ARE-forming mutant SLC4A4 3′UTRs. After 16 h, the transcription was blocked by doxycycline (1 μg/mL) for the indicated periods of time. Total RNA was extracted and subjected RT-QPCR using TaqMan primers specific to Nanouciferase mRNA. The data are presented as luciferase mRNA/RPLPO mRNA levels, mean ± SEM of replicate from one representative experiment with three replicates of at least two experiments. The mRNA half-decay calculations were performed as described in “Methods” using the one-phase exponential decay model

Back to article page