Over-expression of the MBNL protein family members promotes NF1 exon 23a inclusion. (A) Schematic representation of the HMT-NF1 863/499 minigene reporter construct. (B) Co-transfection of CA77 cells with the HMT-NF1 863/499 reporter (0.5 μg) and human MBNL (2 μg) protein expression plasmids. Total RNA was isolated from the transfected cells and semi-quantitative RT-PCR was performed using the primers denoted by arrows in (A). The percentage of NF1 exon 23a inclusion is displayed in the bar graph. Error bars indicate standard deviations and n=3. Total CA77 protein lysate was collected from transfected cells and Western blot analysis was performed using anti-Xpress antibody. Anti-U1 70K was utilized as a loading control.