LiBis: An ultrasensitive alignment method for low-input bisulfite sequencing

Abstract

The cell-free DNA (cfDNA) methylation profile in liquid biopsies has been utilized to diagnose early-stage disease and estimate therapy response. However, in typical clinical settings, only very small amounts of cfDNA can be purified. Whole-genome bisulfite sequencing (WGBS) is the gold standard to measure DNA methylation; however, WGBS using small amounts of fragmented DNA introduces a critical challenge for data analysis, namely a low mapping ratio. This, in turn, generates low sequencing depth and low coverage for CpG sites genome wide. The lack of informative CpGs has become a bottleneck for the clinical application of cfDNA-based WGBS assays. Hence, we developed LiBis (Low-input Bisulfite Sequencing), a novel method for low-input WGBS data alignment. By dynamically clipping initially unmapped reads and remapping clipped fragments, we judiciously rescued those reads and uniquely aligned them to the genome. By substantially increasing the mapping ratio by up to 88%, LiBis dramatically improved the number of informative CpGs and the precision in quantifying the methylation status of individual CpG sites. The high sensitivity and cost effectiveness afforded by LiBis for low-input samples will allow the discovery of genetic and epigenetic features suitable for downstream analysis and biomarker identification using liquid biopsy.Competing Interest StatementThe authors have declared no competing interest.List of AbbreviationsLiBislow-input bisulfite sequencing alignmentcfDNAcell-free DNAWGBSwhole-genome bisulfite sequencingCpGcytosine nucleotide followed by a guanine nucleotidectDNAcirculating tumor DNAHTMLHyperText Markup LanguageHPVHuman papillomavirusView Full Text