GSM5354350: IMR90_NicE-C_rep1; Homo sapiens; OTHER
RNA was harvested using Trizol reagent. Cells were croseelinked with 1% formaldehyde for 10 minutes at room temperature. For Hi-C, we followed in-situ Hi-C protocol, the cells were lysed. Then the lysed cells were digested by MboI overnight at 37℃. DNA fragment ends were filled and labeled with dCTP, dGTP, dTTP and biotin-dATP. Proximity ligation was performed at room temperature for 4 hours. Then the DNA were de-crosslinked at 65°C overnight with Proteinase K and DNA were purified. For NicE-seq, the cells were lysed and then digested by Nt.CviPII and DNA polemerase I (Ecoli)  with dGTP, dTTP, biotin-dATP, biotin-dCTP and 5-methyl-dCTP for 20 min at 37℃. Then the DNA were de-crosslinked at 65°C overnight with Proteinase K and DNA were purified. For NicE-C,  the cells were lysed and then digested by Nt.CviPII and DNA polemerase I (Ecoli)  with dGTP, dTTP, dATP and 5-methyl-dCTP for 20 min at 37℃. DNA fragment ends were filled with dNTP and dA-tailing was performed. Proximity ligation by bridge-linker was performed at room temperature for 4 hours. Then the DNA were de-crosslinked at 65°C overnight with Proteinase K and DNA were purified. Hi-C, NicE-C and NicE-seq DNA were sonicated and binding to M280 beads, then end-repair, dA-tailing and adapter ligation on beads. After PCR amplification and DNA purification, the libraries were sequenced on Illumina NovaSeq 6000 using 150bp paired-end reads. RNA libraries were prepared for sequencing using standard Illumina protocols