Figure 3.

Comparison of Six Different AONs

(A) Schematic drawings show the binding sites of six AONs that bind to different sequences within intron 5 of BBS1. The mutated base pair (red letter) is located in the splice donor site of BBS1 (light green letters). (B) Efficacies to block the intron retention in BBS1 patient-derived fibroblasts were compared between six AONs (AON_1 to AON_6). Therefore, different concentrations of the AONs (9 nM, 18 nM, 36 nM) were incubated for 96 h. Results of the RT-PCR showed that AON_1, AON_2, AON_4, and AON_5 were efficient to block intron retention, whereas AON_3 and AON_6 showed no effect. The size of the different splice products is shown. (C) Control fibroblasts were incubated with AON_1 to AON_6 (36 nM) for 96 h to test for side effects. No splice alteration was detected. NTC, none template control; Patient, fibroblasts homozygous for the BBS1 mutation c.479G > A; Control, fibroblasts with BBS1 reference alleles. CS, correctly spliced transcript; ES, exon skipping; IR, intron retention; kb, kilo bases.