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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Jul 18, 2017 |
| Title |
E95_ElemB |
| Sample type |
SRA |
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| Source name |
E9.5 full embryo
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| Organism |
Mus musculus |
| Characteristics |
strain: Bl6
tissue: full embryo
age: E9.5
genotype: WT
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| Treatment protocol |
4C–seq libraries were constructed as described before (Matelot and Noordermeer, Methods in Molecular Biology 2016). 4C-seq libraries were prepared from 35 E9.5 embryos. 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 1000 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.
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| 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)
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
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| 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.
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| 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
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| Department |
IBENS - UMR ENS - CNRS 8197 - INSERM 1024
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| Street address |
46 Rue d'Ulm
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| City |
Paris |
| ZIP/Postal code |
75005 |
| Country |
France |
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| 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 |
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| Relations |
| BioSample |
SAMN06315571 |
| SRA |
SRX2546901 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2481336_E95_ElemB_4Csignal.bedGraph.gz |
415.1 Kb |
(ftp)(http) |
BEDGRAPH |
| GSM2481336_E95_ElemB_LocalNormalization_11FragSmooth.bedGraph.gz |
101.0 Kb |
(ftp)(http) |
BEDGRAPH |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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