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Sample GSM6956524 Query DataSets for GSM6956524
Status Public on Feb 22, 2023
Title 4Cseq-hg-Patient371-FB-BTRC-Rep2-L21112
Sample type SRA
 
Source name Skin fibroblasts
Organism Homo sapiens
Characteristics cell type: Skin fibroblasts
genotype: Patient with duplication
Extracted molecule genomic DNA
Extraction protocol Skin biopsies were collected from SHFM3 patients and controls by standard procedures. Fibroblasts were cultured in DMEM (Lonza) supplemented with 10% fetal calf serum (Gibco), 1% L-glutamine (Lonza) and 1% penicillin/streptomycin (Lonza).
For 4C-seq libraries fixation and lysis were performed as described for Capture Hi-C. After the first digestion with DpnII (NEB), sticky ends were religated in a 50 ml falcon tube (700 μl 10 ligation buffer (Fermentas), 7 ml H2O, 50 U T4 DNA ligase (Thermo); overnight at 16 °C) and DNA de-cross linked and cleaned as described for the Capture Hi-C. Next, a second digestion (150 μl sample, 50 μl 10× Csp6I buffer (Thermo), 60 U Csp6I (Thermo) 295 μl H2O; overnight at 37 °C) and another re-ligation were performed. For all viewpoints, DNA was purified using a PCR clean up Kit (Qiagen) and 1.6 μg DNA was amplified by PCR (LBX1 Viewpoint: read-primer 5′-TCTATATGCTACCATGATC-3′, secondary-primer 5′-GATGAACTGGAATACCCA-3′; FGF8 Viewpoint: read-primer 5′-AGGGTGCGTTCCAAGATC-3′, secondary-primer 5′-GGTGGCCTGGATGGAAGT-3′; BTRC Viewpoint: read-primer 5′-CAACGCAGCGCCCGGATC-3′, secondary-primer 5′-CTGGGAATGAGGACCTAGGGC-3′). For the library reaction, primers were modified with TruSeq adapters (Illumina): Adapter1 5′-CTACACGACGCTCTTCCGATCT-3′ and Adapter2 5′-CAGAC GTGTGCTCTTCCGATCT-3′. Between 50 and 200 ng were used as input of a single 4C PCR reaction depending on the complexity. The reaction was performed in a 50 μl volume using the Expand Long Template System (Roche) and 29 reaction cycles. After the PCR all reactions were combined and the DNA purified with a PCR clean up Kit (Qiagen).
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina NovaSeq 6000
 
Data processing Sequencing reads were mapped, normalized and smoothed with pipe4C53 using the reference genome GRCh37 and default settings. All viewpoints were performed in replicates and as quality measure >70% of reads were mapped within a size range of 1Mb and >80% within 100kb around the viewpoint.
Assembly: hg19
Library strategy: 4C-seq
 
Submission date Jan 22, 2023
Last update date Feb 23, 2023
Contact name Giulia Cova
E-mail(s) Giulia.cova@nyulangone.org
Organization name Max Planck Institute
Street address 63-73 Ihnestrasse
City Berlin
ZIP/Postal code 14195
Country Germany
 
Platform ID GPL24676
Series (2)
GSE197404 Combinatorial effects on gene expression at the Lbx1/Fgf8 locus resolve Split-Hand/Foot Malformation type 3
GSE223454 Combinatorial effects on gene expression at the Lbx1/Fgf8 locus resolve Split-Hand/Foot Malformation type 3 [4C-seq]
Relations
BioSample SAMN32868381
SRA SRX19248032

Supplementary file Size Download File type/resource
GSM6956524_4Cseq-hg-Patient371-FB-BTRC-Rep2-L21112.bedgraph.gz 3.2 Mb (ftp)(http) BEDGRAPH
GSM6956524_4Cseq-hg-Patient371-FB-BTRC-Rep2-L21112.wig.gz 1.5 Mb (ftp)(http) WIG
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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