GSM9040595: Input-1; Homo sapiens; OTHER
NB4 cell lysates were prepared from 20x10 to6 cells each, cross-linked with 1% glutaraldehyde and sonicated. MALNC was captured with 30 biotinylated tilling probes (Thermo Scientific) which were designed using Biosearch Technologies' Stellaris FISH Probe Designer. Probes were divided into an odd and an even probe pool (15 probes each) and separately subject to hybridization with cross-linked and sonicated chromatin. MALNC-chromatin complexes were pulled down using streptavidin magnetic beads (Thermo Scientific). MALNC pull-down was confirmed by RNA extraction followed by qRT-PCR detection of MALNC expression. Probes against LacZ were used as negative control (Magna ChIRP™ Negative Control Probe Set, Merck) accounting for non-specific binding. gDNA was extracted (Qiagen DNeasy Blood & Tissue Kit) and subject to DNA sequencing. Sequencing libraries were prepared from the entire sample volume of ChIRP-DNA using the Smarter ThruPLEX DNA- seq Kit without Covaris fragmentation (cat# R400676, Takara) with Unique dual index Set A-D (cat# R400665/6/7/8, Takara). The library preparation was performed according to the manufacturers' instructions (guide#112219). Paired-end 150bp read length, NovaSeq 6000 system, SP flowcell and v1.5 sequencing chemistry. A sequencing library for the phage PhiX was included as 5% spike-in in the sequencing run.