NGS-based deep bisulfite sequencing

We have developed an NGS-based deep bisulfite sequencing protocol for the DNA methylation analysis of genomes. This approach allows the rapid and efficient construction of NGS-ready libraries with a large number of PCR products that have been individually amplified from bisulfite-converted DNA. This approach also employs a bioinformatics strategy to sort the raw sequence reads generated from NGS platforms and subsequently to derive DNA methylation levels for individual loci. The results demonstrated that this NGS-based deep bisulfite sequencing approach provide not only DNA methylation levels but also informative DNA methylation patterns that have not been seen through other existing methods. • This protocol provides an efficient method generating NGS-ready libraries from individually amplified PCR products. • This protocol provides a bioinformatics strategy sorting NGS-derived raw sequence reads. • This protocol provides deep bisulfite sequencing results that can measure DNA methylation levels and patterns of individual loci.