Figure 2: SNRPB mutations in the alternative (alt) PTC-containing exon cause increased exon inclusion.
From: Disrupted auto-regulation of the spliceosomal gene SNRPB causes cerebro–costo–mandibular syndrome
(a,b) Cloning of the alternative exon with either the chr20:g.2447951C>G (mut1) or chr20:g.2447847G>T (mut2) mutation into a splicing minigene reporter and transfection into HEK297 cells shows 78% and 80% exon inclusion, respectively, compared with 23% for the wild-type (WT) sequence. Results shown are from one representative experimental replicate of three. A 100-bp DNA ladder was used as a size marker in a. NT, no template control. (c) Patient fibroblasts with the chr20:g.2447951C>G (pt 1), chr20:g.2447952C>G (pt 2) and chr20:g.2447847G>T (pt 3) mutations show increased expression of the PTC-containing transcript by qRT–PCR. (d) The same three patients show decreased total SNRPB expression by qRT–PCR. In c,d, the grey columns represent the normalized average expression from three anonymous controls (Ctrl). The experiment was performed three times. For b–d, statistical significance was determined with a Student’s t-test. Bars indicate s.d. * indicates 0.005<P<0.05, ** indicates 0.0005<P<0.005 with *** indicating P<0.001 and ****P<0.0001.
