The expression level of reprogramming factors could affect the outcome of reprogramming (Sommer et al., 2012) and lead in part to the heterogeneity of NGN2-iNs. We thus analyzed the relationship between NGN2 expression and the molecular identity of corresponding NGN2-iNs. The NGN2 expression level and proportion of NGN2-expressing cells was indeed lower in the off-target cluster (CD99+), in line with previous studies that failed reprogramming is linked to silenced reprogramming factors (Treutlein et al., 2016) (Figure 3F). Among the successfully reprogrammed neural clusters, we observed variable expression levels of NGN2 and proportion of NGN2-expressing cells, prompting us to examine whether NGN2 dosage affects the NGN2-iN reprogramming heterogeneity.
