GSM7520429: ChIP-seq of H3K4me1 in Homo sapiens: U87-H3K4me1-Rep1; Homo sapiens; ChIP-Seq
1. For RNA-seq: cells were lysed with Trizol reagent (Life Technologies) and RNA was extracted according to the manufacturer's instructions.  2. For Chip-seq: Cells were cross-linked with 1% formaldehyde at RT for 10 min and quenched in 125 mM glycine for 5 min. Cells were resuspended in SDS lysis buffer for 5 min and sonicated to generate DNA fragments averaging 100–500 bp in length. After centrifugation, chromatin fragments were immunoprecipitated with antibodies overnight (H3K27ac, H3K4me1, Abcam). The precipitated DNA and input DNA were purified and then sequenced at Novogene (Tianjin city, China).                                                                                                                                                                                                                                            3. For HiChIP: Briefly,  cells were cross-linked and chromatin was digested using MboI restriction enzyme (NEB). Then biotin-14-dATP was used to fill in the restriction fragment overhangs and mark the DNA ends. After ligation and sonication, the genomic DNA was incubated with H3K27ac antibody at 4 °C overnight. Immunocomplexes were captured by protein A magnetic beads. Beads were subsequently washed three times with low salt wash buffer, high salt wash buffer and LiCl wash buffer. DNA was eluted and purified with AMPure XP Beads (Beckman). Biotinylated DNA beads were captured by Streptavidin C-1. QIAseq FX DNA Library Kits (QIAGEN) was used to generate the sequencing library according to the manufacturer's protocol. The libraries were validated for size distribution of 300–700 bp using AMPure XP beads (Beckman). The DNA was subjected to 2 × 150 bp paired-end sequencing at Novogene (Tianjin city, China). RNA-seq Library: Total RNA was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer's recommendations and index codes were added to attribute sequences to each sample. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in NEBNext First Strand Synthesis Reaction Buffer(5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase (RNase H). Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3' ends of DNA fragments, NEBNext Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 250~300 bp in length, the library fragments were purified with AMPure XP system. Then 3 µl USER Enzyme (NEB, USA) was used with size selected, adaptor-ligated cDNA at 37°C for 15 min followed by 5 min at 95 °C before PCR. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system. The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumia) according to the manufacturer's instructions. After cluster generation, the library preparations were sequenced on an Illumina Novaseq 6000 platform and 150 bp paired-end reads were generated.                                                                                                                                                         2. ChIP-seq:  DNA libraries were generated using a FS DNA Library Prep Kit (QIAGEN). The precipitated DNA and input DNA were purified and then sequenced on Illumina HiSeq XTen sequencer. Bowtie2 was used to map ChIP-seq raw reads to the hg38 human reference genome. MACS2 program was used to call peaks of ChIP-seq data with the corresponding input data as control with default parameters. 3. HiChIP: QIAseq FX DNA Library Kits (QIAGEN) was used to generate the sequencing library according to the manufacturer's protocol. The libraries were validated for size distribution of 300–700 bp using AMPure XP beads (Beckman). The DNA was subjected to 2 × 150 bp paired-end sequencing. The respective paired-end reads were aligned to the hg38 genome by HiC-Pro pipeline. Default settings were used for removal of duplicated reads, assignment to MboI restriction fragments and filtering for valid interactions. For each cell type, we merged the valid read pairs of individual samples. Hichipper was applied to call loops with default parameter. Loops with FDR < 0.1 and supported by at least 2 paired-end tags (PETS) were kept for further analysis. HiC-Pro was used to generate the HiChIP interaction maps. HiChIP interaction maps was then visualized by JuicerBox software at 50, 10 and 5 kb resolution as indicated.