Well, from your table S5 and table S12, as is usual we see that about 0.1 - 1% of the cells were mutants in the control group (ie they came pre-"modified"). In

Well, from your table S5 and table S12, as is usual we see that about 0.1 - 1% of the cells were mutants in the control group (ie they came pre-"modified"). Interestingly, from table S12 and my (selection for pre-existing mutants) model we can also explain a mysterious result they observed: "Silent co-mutations in the repair oligonucleotides were introduced into >99% of D54H mutant alleles and ~3% of the F482S allele (Fig. 2c). Carryover of these silent SNPs indicates that the alleles are the product of HR and not de novo mutagenesis. The lower rate of coappearance of silent SNPs in F482S is presumably due to the larger distance between the two SNPs in the oligonucleotide and is rather common to see with single-stranded oligonucleotide donors19." Rather than that ad hoc explanation, it is simply that the D54H cells had more silent only mutations to begin with (0.1% vs 0.0%). Regarding that 0.0%, an annoying thing is that they only report these percentages to one decimal place. I'm not exactly clear on the number of cells present before the CRISPR-Cas9 treatment, but it sounds like 10^8, and then they let them grow for 72 hr + 7-12 days (total of 10-15 days) after the treatment. They also don't tell us how many cells were left at the end... but anyway if we assume these cells divide once a day, and 0.1% are preexisting mutants we could calculate the possible number of mutants thus: Nt = 10^8 p = 0.001 d = 0:15 Nt*p*2^d After 12 days we can get ~400 million cells from those initial pre-existing mutants, and after 15 days over as 3 billion. Of course other factors would probably come into play that limit this growth, I'm just saying it would be no problem for that small subset of the population to become dominant during the experiment. That is even if the 99,900,000 "WT" cells were just growth arrested rather than died. So I find those results to favor the "selection for pre-existing mutants" explanation over the "gene modification" one.