Decoding Sanger Sequencing Chromatograms from CRISPR-Induced Mutations.

In many diploid organisms, the majority mutations induced by clustered regularly interspaced short palindromic repeats (CRISPR)-mediated genome editing are non- chimeric, including biallelic, homozygous, and heterozygous mutations. Direct Sanger sequencing of the PCR amplicons containing non-homozygous mutations superimposes sequencing chromatograms, displaying overlapping peaks beginning from the mutation sites. In this chapter we describe the degenerate sequence decoding (DSD) strategy and its automatic web-based tool, DSDecodeM, for decoding the Sanger sequencing chromatograms from different types of targeted mutations. DSDecodeM, as a convenient and versatile tool, can considerably facilitate the genotyping work of CRISPR-induced mutants.