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. 2019 Oct 31;20(21):5434. doi: 10.3390/ijms20215434

Figure 3.

Figure 3

Comparison of ATXN3 exon 10 skipping in patient cells, transfected with 2′-Me PS AOs and PMOs. Cells were harvested 48 hr following transfection for protein and RNA analysis. Agarose gel fractionation of ATXN3 amplicons, following 2′-Me PS AO (A) and PMO (B) cocktail transfections at concentrations of 400 nM and 20 μM, respectively shows full-length (FL 74Q; FL 24Q) and induced transcript products after skipping of exon 10 (Δ10). Ataxin-3 protein was analysed by Western blotting following 2′-Me PS AO (C) and PMO (D) cocktail transfection at a concentration of 400 nM and 20 μM, respectively. The disease-causing 74Q protein is approximately 60 kDa, the protein encoded by the healthy allele is approximately 48 kDa and the Δ10 encoded protein, 34 kDa. Beta-Tubulin was used as a loading control. The samples were also probed with an anti-polyglutamine antibody to identify the pathogenic stretch of glutamines in the ataxin-3 protein. Densitometric analysis performed on the Western blots are shown below the blots (means plus error bars. Error bars = standard deviation, n = 3). Samples are normalised to the GT control. (FL = full-length, Δ10 = exon skipped product, GT = Gene Tools control PMO, Q = glutamine, PolyQ = polyglutamine).