Analysis of Dystrophin Isoforms Induced by PPMO-Mediated Exon Skipping In Vitro
Single-round and nested RT-PCR analysis of DMD transcripts confirming robust skipping of exon 23, exons 56+57, 58+59, and exon 70 in vitro. An RT-PCR no-template negative control was loaded in the final lane. Transcript product size in base pairs (bp) are indicated by 100 bp DNA ladder. (A) The level of exon 23 skipping after individual exon 23 PPMO treatment. (B) Analysis of exon 23 skipping after treatment with PPMO cocktails targeting exons 23+56+57 and 23+58+59. (C) Analysis of exon 23 skipping after exon 23+70 exon skipping PPMO cocktail treatment. (D) Exons 56+57 and 58+59 skipping after PPMO cocktail treatment. (E) Exon 70 skipping after treatment with the AO cocktail targeting exons 23+70. Cocktail 1, a combination of PPMOs targeting exon 23+58+59 skipping; cocktail 2, a combination of PPMOs to skip exons 23+56+57; cocktail 3, a preparation of PPMOs to skip exons 23+70. FL, full-length amplicon; Δ23, exon 23 skipped amplicon; Δ56+57, exons 56+57 excised amplicon; Δ58+59, exons 58+59 removed amplicon; Δ70, exon 70 skipped amplicon; GTC, gene-tools control PPMO; UT, untreated; -Ve, no template negative control.