All samples were displayed on a scatter plot, with the number of sequenced molecules on the x-axis and number of variant molecules on the y-axis (Fig 3). Variant allele frequencies are indicated by broken diagonal lines. The variant allele frequency of filtered and removed variants ranged from 0.25–76.1% and from 0.35–45.9%, respectively. However, the distribution patterns between the two types of variants were different. The variant allele frequency of filtered variants varied widely in their respective proportions, from more than 10% to less than 1% of sequenced molecules. In contrast, the removed variants rarely made up more than 10% of all sequenced molecules, and most were approximately 1%. More than two-thirds of variants isolated from tissue samples showed high variant allele frequency.