Sequencing of DNA fragments with 5-hydroxymethylcytosine modification obtained from cell-free DNA of patients diagnosed with osteosarcoma
Nano-hmC-Seal libraries were constructed from 1-10 ng of  cell-free (cfDNA). Briefly, after ligation with sequencing adapters using the KAPA Hyper Prep Kit (KAPA Biosciences), cfDNA 5-hmC marks were subjected to T4 beta-glucosyltransferase enzymatic modification with UDP-N3-glucose, followed by subsequent chemical modification with biotin-PEG4-dibenzocyclooctyne (DBCO). Biotin-labeled cfDNA fragments were then pulled down using streptavidin M270 Dynabeads (Invitrogen), and PCR amplified to construct sequencing libraries which were purified with AMPure XP beads (Beckman Coulter) to construct libraries. Fifty base-pair, paired-end libraries were sequenced.
NCI-phs003842
16_1