For suppression of miR-34a expression, hNPCs (unaffected male, derived from GM08330 iPSCs) were transfected after 12 days of differentiation with either a construct expressing an anti-miR-34a sequence (GeneCopoeia, HmiR-AN0440-AM03-B) or a control scrambled sequence (GeneCopoeia, CmiR-AN0001-AM03). Since both constructs co-expressed mCherry as a reporter gene, for these experiments the 2 days transfected cells were fixed cells and immunostained for mCherry to allow complete visualization of the cells expressing the anti-miR constructs. The Sholl analysis was done as described above for the miR-34a overexpressing hNPCs. To assess the effect of the anti-miR-34a expression on target mRNAs, transfected cells were selected for 2 days using an antibiotic selection with hygromycin B at 200µg/ml.
