The initial bisulfite treatment is gentle on cfDNA yet effective in converting any unmodified cytosines into uracil. Next, the innovative splinted adapters capture and directly ligate the Illumina-compatible adapters onto any size DNA fragment, allowing for sequencing of nicked, damaged, and short DNA fragments that may have otherwise been discarded with traditional library prep methods. The direct ligation of the adapters also eliminates the need for second strand synthesis, end repair, and dA tailing steps, thus reducing bias by preserving the integrity of any native methylation that is present on the fragment termini. Finally, the adapter-ligated cfDNA is indexed and amplified via PCR with Zymo-Seq UDIs, generating libraries that are ready for sequencing on any Illumina instrument.
HIGHLIGHTS:
·Optimized for small fragment input: Ideal for small and damaged DNA fragments such as cell-free DNA (cfDNA).
·Accurate methylation calling: Direct ligation-based protocol allows for accurate reads and methylation calling of native termini for each DNA fragment.
·Streamlined and simple workflow: Prepare robust methyl-seq libraries in as little as 3 steps.