GEO Accession viewer

NCBI > GEO > Accession Display

Not logged in | Login

GEO help: Mouse over screen elements for information.

Sample GSM4340726

Query DataSets for GSM4340726

Status

Public on Aug 13, 2020

Title

Hi-C Msc Experiment #2

Sample type

SRA

Source name

BM-hMSC-TERT4

Organism

Homo sapiens

Characteristics

cell type: Stem cell
Stage: Day 0

Treatment protocol

Two days post confluency (day 0), cells were induced to undergo adipocyte differentiation by exposing them to a adipogenic differentiation cocktail (DMEM supplemented with 10% fetal calf serum, 10ug/mL insulin, 1µM rosiglitazone, 100 mM dexamethasone, and 500 µM isobutylmethylxanthine). Medium was replaced on days 2, 4, 7 and 9.

Growth protocol

Telomerase-immortalized human mesenchymal stromal cells of bone marrow (BM-hMSC-TERT4) were grown under standard cell culture conditions in α-MEM supplemented with 10 % fetal calf serum (FCS) and 1 % penicillin/streptomycin (P/S).

Extracted molecule

genomic DNA

Extraction protocol

Hi-C was performed using in-nucleus ligation following a modified version of a previously described protocol (Nagano et al., 2015). Chromatin was digested using MboI restriction enzyme. The digested ends were filled in using biotinylated d-ATP and ligated. Chromatin was de-crosslinked and purified by phenol-chloroform extraction. Purified DNA (50 µg) was sheared to an average size of 400 bp by sonication (Covaris). The sheared DNA was end-repaired, adenine-tailed and double size-selected using AMPure XP beads to isolate DNA ranging from 250 to 550 bp. Ligation fragments marked by biotin were immobilized using MyOne Streptavidin C1 DynaBeads (Invitrogen) and ligated to NEBNext hairpin adaptors (New England Biolabs) followed by U excision by USER Enzyme (New England Biolabs). The resulting Hi-C libraries were amplified using universal primer and 6bp-index primers (New England Biolabs).
Hi-C libraries were constructed from genomic DNA according to the manufacturer's instructions (Illumina).

Library strategy

Hi-C

Library source

genomic

Library selection

other

Instrument model

Illumina HiSeq 1500

Description

HiC_Msc.bed.gz

Data processing

Base calling for all sequencing runs was performed using bcl2fastq (v2.20.0.422).
ChIP-seq: Reads were aligned using STAR (v2.3.1, --outSJfilterIntronMaxVsReadN 0 --outFilterMatchNmin 25 --outFilterMismatchNmax 2 --alignIntronMax 1). Duplicated reads/fragments were removed using Picard (v2.5.0). Reads/fragments were counted using HOMER (v4.10.3) in DNase I hypersensitive regions identified in a previous study (Rauch et al., 2019)
scATAC-seq: Reads (R1 and R2) were trimmed using TrimGalore (v. 0.6.0) and aligned aligned using STAR (v2.3.1, --outSJfilterIntronMaxVsReadN 0 --outFilterMatchNmin 25 --outFilterMismatchNmax 2 --alignIntronMax 1). Only short fragments (< 140bp) were kept , the short fragments were deduplicated using Picard (v2.5.0) and peaks were detected using MACS2 (v. 2.1.2). The cell barcodes (Index1) were quality filtered using FASTX toolkit (v. 0.0.13, -q 30 -p 80). The quality filtered barcodes were matched to the barcode white list (10X Genomics) and only barcodes with more than 10000 reads were kept. The raw aligned reads were split into cell-specific files using the filtered barcodes and individually deduplicated. Count matrices were generated by counting fragments for each cell using featureCounts (v.2.0.0) in peak detected regions. Cells and peaks were quality filtered and co-accessible peaks were identified using Cicero.
CUT&RUN: Reads trimmed with cutadpt and aligned using STAR (v2.3.1, --outSJfilterIntronMaxVsReadN 0 --outFilterMatchNmin 25 --outFilterMismatchNmax 2 --alignIntronMax 1).
RNA-seq: Reads were aligned using STAR (v2.3.1). Reads were counted using iRNA-seq (v1.0).
HiC: Reads were aligned and quality controlled using Hicup (v0.6.1). TADs were identified using HiTAD from TADlib (v0.3.1).
ECHiC: Reads were aligned and quality controlled using Hicup (v0.6.1). Interactions between virtual restriction fragments were detected using Chicago (v1.6.0) using custom weights calculated from high confidence interactions in the data.
Genome_build: hg19
Supplementary_files_format_and_content: ChIP-seq: Tab-delimited txt files containing normalized tag counts for each replicate in DNase I hypersensitive regions identified in a previous study (Rauch et al., 2019). For each replicate, read depth normalized bedGraph files are also supplied for filtered and deduplicated alignments.
Supplementary_files_format_and_content: scATAC-seq: Tab-deliminted txt file containing the peak co-accessibility (estimated with Cicero).
Supplementary_files_format_and_content: CUT&RUN: Tab-delimited txt files containing raw tag counts for each replicate in DNase I hypersensitive regions identified in a previous study (Rauch et al., 2019). For each replicate, read depth normalized bedGraph files are also supplied.
Supplementary_files_format_and_content: RNA-seq: Tab-delimited txt file containing raw tag counts for each replicate for all RefSeq genes . For each replicate, read depth normalized bedGraph files are also supplied.
Supplementary_files_format_and_content: Hi-C: The BED file contains all detected TADs and subTADs (identified using HiTAD).
Supplementary_files_format_and_content: ECHi-C: All interactions identified (using Chicago), as well as counts in all samples and annotation is also supplied in a tab-delimited txt file.

Submission date

Feb 26, 2020

Last update date

Aug 13, 2020

Contact name

Susanne Mandrup

E-mail(s)

s.mandrup@bmb.sdu.dk

Phone

+45 6550 2340

Organization name

University of Southern Denmark