title: GSM3182431: HH30_PHL_WT_Hoxd12vp; Gallus gallus; OTHER
library_construction_protocol: The dissected limb bud tissues were dissociated with collagenase (Sigma Aldrich) and filtered through cell strainer to obtain single cells. Cells were fixed with 2% formaldehyde for 10 min at room temperature and the reaction was quenched on ice with glycine. Cells were further lysed with 10 mM Tris pH 7.5, 10 mM NaCl, 5 mM MgCl2, 0.1 mM EGTA, 1x Protease inhibitor cocktail to isolate nuclei and stored at -80°C. Nuclei from pools of samples were digested with NlaIII (New England Biolabs) and ligated with T4 DNA ligase HC (Promega) in diluted conditions to promote intramolecular ligation. Sequences ligated to fragments of interest were digested again with DpnII (New England Biolabs), ligated with T4 DNA ligase HC (Promega) in diluted conditions. These templates were amplified using Expand long template (Roche) and inversed PCR primers flanked with adaptors allowing multiplexing. All primer and barcode information are described in Supplemental Table 3 in Yakushiji-Kaminatsui et al. (2018).