Each duplex sequencing read contains a fixed 5-nucleotide sequence (shown in figures in black) located upstream of the 12-nucleotide tag sequence. The reads are filtered if they do not have the expected 5-nucleotide sequence or have more than nine identical or ambiguous bases within each tag. The two 12-nucleotide tags at each end of the reads are combined and moved to the read header. Two families of reads are formed that originate from the two strands of DNA. One family contains reads with αβ header originating from strand 1 and the second contains reads with βα header originating from strand 2 (Figure 2). The reads are then trimmed by removing the fixed 5-base pair sequence and 4 error-prone nucleotides located at the sites of ligation and end repair. The remaining reads are assembled to consensus sequences using SSCS and DCS assemblies.