COVID-19 Whole-Genome Resequencing with Redundant Tiling PCR and Subtract-Based Amplicon Normalization Successfully Characterized SARS-CoV-2 Variants in Clinical Specimens

Figure 3

Subtraction of biotinylated amplicons successfully reduced the number of biotinylated amplicons. (a) The NIID, BGD1, 2, and 3 were resequenced with or without biotinylated amplicon subtraction. The read counts for each amplicon region were normalized from 100,000 mapped reads and were log10-transformed. Amplicons with biotinylated primers (blue: biotinylated) and without biotinylated primers (red: normal) are shown for each sample. (b) Fold changes of normalized amplicon counts for each amplicon from the control and subtracted experiments are shown. Distributions of the fold changes are shown in the violin plots, and the 1st quantile, median, and 3^rd quantile values are shown in the box plots. Amplicons with biotinylated primers (biotinylated) and without biotinylated primers (normal) are shown for each sample. (c) Normalized read depths for each sample with or without subtraction are shown. The regions with low depth, genome positions 3000–3500 bp (left panel) and 8000–8500 bp (right panel) are also shown. Solid lines: depths for the experiments with subtraction (subtract). Dashed lines: depths for the experiments without subtraction (ct).

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