Because the ATM western blot (Figure 1D) appears to show different quantities of ATM protein at different passage numbers of Q3SC, we wondered if early passage cells might have a different genotype from later passage cells. Early passage (P4) DNA prepared from Q3SC exhibited c.217_218 delGA in one allele, matching Q3SA and the diagnostic genotype for the JHU_Q3 subject, but later passage cells (P10) lacked this mutation (Figure 2B). This suggests that the Q3SC iPSC culture may have acquired a spontaneous gene reversion during early passaging. Alternatively, this culture could have suffered from cross-contamination. The latter seems unlikely, because the ATM mutation genotype for Q3SC does not match the maternal heterozygous carrier that we had thawed and reprogrammed (CAR3) but instead matches the imputed genotype and gender of cells from the father (CAR4), from whom we stored blood samples that have never been thawed or cultured in our laboratory.
