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Sample GSM2481353 Query DataSets for GSM2481353
Status Public on Jul 18, 2017
Title E85H_ElemB
Sample type SRA
 
Source name E8.5 head
Organism Mus musculus
Characteristics strain: Bl6
tissue: head
age: E8.5
genotype: WT
Treatment protocol 4C–seq libraries were constructed as described before (Matelot and Noordermeer, Methods in Molecular Biology 2016). 4C-seq libraries were prepared from 250 micro-dissected E8.5 embryonal heads. The primary restriction enzyme used was DpnII (New England Biolabs, R0125L) and the secondary restriction enzyme was NlaIII (New England Biolabs, R0543M). For each viewpoint 1400 ng of the 4C-seq library was amplified using 16 individual PCR reactions with inverse primers containing Illumina Solexa adapter sequences.
Circular Chromosome Conformation Capture (4C) technique was performed as described (Matelot and Noordermeer, Methods in Molecular Biology 2016). Cross-linked chromatin was digested with DpnII, ligated under diluted conditions, cross links were reversed and DNA was further digested by NlaIII and again ligated under diluted conditions.
Extracted molecule genomic DNA
Extraction protocol Multiplexed samples were sequenced on the Illumina Genome HiSeq 2500 system using 100 bp single-end reads according to the manufacturer’s specifications. 4C-seq reads were sorted, aligned, and translated to restriction fragments using the 4C-seq pipeline of the BBCF HTSstation (David et al. 2014, Plos ONE and available at http://htsstation.epfl.ch). Samples were mapped to the ENSEMBL Mouse assembly NCBIM37 (mm9).
Illumina adapters (single end reads) were included in the inverse 4C PCR primers used for 4C-seq library preparation
Cells were isolated in PBS supplemented with 10% Fetal Calf Serum. Tissue sample were further made single cell by collagenase treatment and applying a cell-strainer cap (BD Biosciences). Chromatin was crosslinked in the presence of 2% formaldehyde.
PCR amplification using primers with included adapter sequences (for Illumina HiSeq2500)
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina HiSeq 2500
 
Data processing Library strategy: 4C-Seq
De-multiplexing, mapping and 4C-seq were done through HTSstation (David et al, 2014 PLoS ONE and http://htsstation.epfl.ch/).
Reads were mapped to the mm9 (NCBIM37) Mouse genome assembly with bowtie version 0.12.7 (Langmead et al, 2009 Genome Biology) with parameters -n 2 --best -strata -m 5.
Mapped reads were translated to restriction fragments using the HTSstation (David et al, 2014 PLoS ONE and http://htsstation.epfl.ch/).
4C figures were made using a running mean algorithm with a window size of eleven fragments.
Genome_build: mm9
Supplementary_files_format_and_content: Signal/restriction fragment (bedGraph) and locally normalized, smoothed (bedGraph), one per viewpoint, per cell type.
 
Submission date Feb 09, 2017
Last update date May 15, 2019
Contact name Pascale Gilardi-Hebenstreit
Organization name Institut de Biologie de l'Ecole Normale Superieure
Department IBENS - UMR ENS - CNRS 8197 - INSERM 1024
Street address 46 Rue d'Ulm
City Paris
ZIP/Postal code 75005
Country France
 
Platform ID GPL17021
Series (2)
GSE94715 Krox20 hindbrain regulation involves multiple modes of cooperation between cis-acting elements [4C-seq]
GSE94716 Krox20 hindbrain regulation involves multiple modes of cooperation between cis-acting elements
Relations
BioSample SAMN06315554
SRA SRX2546918

Supplementary file Size Download File type/resource
GSM2481353_E85H_ElemB_4Csignal.bedGraph.gz 296.0 Kb (ftp)(http) BEDGRAPH
GSM2481353_E85H_ElemB_LocalNormalization_11FragSmooth.bedGraph.gz 98.5 Kb (ftp)(http) BEDGRAPH
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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