Extended Data Fig. 2: Quality control of colonies from single-cell derived clones.

Example histograms of the variant allele fraction (VAF—the proportion of sequencing reads that report the mutation) of mutations in single colonies. a, The VAF of all mutations on autosomes in a typical clonal colony. Because there are two copies of each autosome, and each mutation occurs on only one of them, in a clonal sample the VAF of autosomal mutations is binomially distributed with a mean of 0.5. b, The VAF of all mutations on the X chromosome in the same clonal colony. Because the subject is male, there is only one copy of the X chromosome, and so true mutations here must have a VAF of 1. Occasionally, lower VAFs are seen when a mutation is not detected on a read, when a read from another locus is aberrantly mapped to the locus in question and thus lowering the apparent coverage, or when a mutation is acquired in vitro. c, d, The VAF of autosomal and X chromosome mutations, respectively, in a typical colony seeded by more than one cell. As not all the reads come from the same cell, and most mutations are private to a given cell, a lower proportion of DNA molecules carry the mutation in a polyclonal colony than in a clonal colony, resulting in a leftward shift of the peak of the VAF histogram. These histograms suggest that the number of mutations acquired by the colonies after a few weeks of in vitro expansion is a small fraction of those acquired in vivo over 60 years of life.