HIC seq of Kmeria septentrionalis: adult leaf
Briefly, the 50 ng genomic DNA was fixed in situ in the nucleus using formaldehyde, resulting in cross-linking of DNA that is in close spatial proximity.The cross-linked DNA was then digested with the restriction enzyme provided in the kit. The digested ends were filled in with biotinylated nucleotides to label the DNA ends at the cross-linked junctions. Subsequent ligation of the blunt-end fragments which are in close proximity in 3D space was performed under dilute conditions to favor intramolecular ligation.After ligation, cross-links were reversed and the DNA was purified from proteins and other cellular debris. The resulting DNA, now enriched for biotin-labeled ligation junctions, was sheared to a length suitable for sequencing using a focused-ultrasonicator (Covaris). Biotinylated DNA fragments were then pulled down with streptavidin beads, followed by library preparation steps such as end repair, A-tailing, and adapter ligation as per the kit's guidelines.