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Sample GSM786046 Query DataSets for GSM786046
Status Public on Nov 28, 2011
Title Limb 4C isIV
Sample type genomic
 
Channel 1
Source name Island IV interacting sequences from E12.5 mouse Forelimbs/autopods
Organism Mus musculus
Characteristics age: E12.5
developmental stage: embryo
tissue: Forelimb/ autopod
viewpoint: IslandIV
genetic background: B6/CBA (F1)
Extracted molecule genomic DNA
Extraction protocol Presumptive digits and forebrains were dissected from E12.5 mouse embryos, dissociated by collagenase treatment and fixed in 2 % formaldehyde for 10 minutes at room temperature. After cell lysis, nuclei were stored at -80°C. Pools of 16 autopods or 2 forebrains were digested with DpnII (New England Biolabs) and ligated in diluted conditions to promote intramolecular ligation events. DNA was purified by phenol/chloroform extraction followed by ethanol precipitation. Sequences ligated to the fragment of interest (interacting regions) were amplified by inverse PCR as described (Simonis et al., 2006), using the following primer pairs for each viewpoint: Hoxd13-F: 5'-GCACAGGCTCTTCGTCGT-3'; Hoxd13-R: 5'-CAAATTCGAGACAGCTCATCC-3'; Hoxd4-F: 5'-GTAGTTCAGAGCGGGGTTTG-3'; Hoxd4-R: 5'-GGCAGCTGGGATTAACACTTA-3'; IslandI-F: 5'-GGGACACGGTCCAATTTAAG-3'; IslandI-R: 5'-TGGAAAGGTCATGCGTGTT-3'; IslandIV-F: 5'-GGAATGACCAAGGGATACAAGA-3'; IslandIV-R: 5'-GGGTAAAGACTGCACAAGAGC-3'; Prox-F: 5'-GACGGCCACTCACATAATAAGA-3'; Prox-R: 5'-GGCTTGACAGCTGTGTAGTAGC-3'.. 200 ng of 4C template were amplified per reaction, using AmpliTaq DNA polymerase (Applied Biosystems). For each condition, 16 reactions were pooled and purified using Qiagen PCR clean-up kit.
Label biotin
Label protocol 1 ug of 4C and input DNA was fragmented and labeled using GeneChip WT Double-Stranded DNA Terminal Labeling Kit (Affymetrix).
 
Channel 2
Source name Input DNA from E12.5 mouse limbs
Organism Mus musculus
Characteristics age: E12.5
developmental stage: embryo
tissue: Forelimb/ autopod
genetic background: B6/CBA (F1)
Extracted molecule genomic DNA
Extraction protocol Presumptive digits and forebrains were dissected from E12.5 mouse embryos, dissociated by collagenase treatment and fixed in 2 % formaldehyde for 10 minutes at room temperature. After cell lysis, nuclei were stored at -80°C. Pools of 16 autopods or 2 forebrains were digested with DpnII (New England Biolabs) and ligated in diluted conditions to promote intramolecular ligation events. DNA was purified by phenol/chloroform extraction followed by ethanol precipitation. Sequences ligated to the fragment of interest (interacting regions) were amplified by inverse PCR as described (Simonis et al., 2006), using the following primer pairs for each viewpoint: Hoxd13-F: 5'-GCACAGGCTCTTCGTCGT-3'; Hoxd13-R: 5'-CAAATTCGAGACAGCTCATCC-3'; Hoxd4-F: 5'-GTAGTTCAGAGCGGGGTTTG-3'; Hoxd4-R: 5'-GGCAGCTGGGATTAACACTTA-3'; IslandI-F: 5'-GGGACACGGTCCAATTTAAG-3'; IslandI-R: 5'-TGGAAAGGTCATGCGTGTT-3'; IslandIV-F: 5'-GGAATGACCAAGGGATACAAGA-3'; IslandIV-R: 5'-GGGTAAAGACTGCACAAGAGC-3'; Prox-F: 5'-GACGGCCACTCACATAATAAGA-3'; Prox-R: 5'-GGCTTGACAGCTGTGTAGTAGC-3'.. 200 ng of 4C template were amplified per reaction, using AmpliTaq DNA polymerase (Applied Biosystems). For each condition, 16 reactions were pooled and purified using Qiagen PCR clean-up kit.
Label biotin
Label protocol 1 ug of 4C and input DNA was fragmented and labeled using GeneChip WT Double-Stranded DNA Terminal Labeling Kit (Affymetrix).
 
 
Hybridization protocol 1μg of DNA was hybridzed per array for 16 hours at 45° at 60rpm using the Affymetrix hybridization kit.
Scan protocol Arrays were scanned on an Affymetrix Scanner 3000 7G
Description Island IV interacting sequences from E12.5 mouse Forelimbs/autopods, Biological Rep. 1+2
Data processing Data were quantile normalized within 4C/input replicate groups and scaled to medial feature intensity of 100 using TAS software (Affymetrix). For each genomic position, a data set was generated consisting of all (PM-MM) pairs mapping within a sliding window of 250 bp.
signal.bar and pvalue.bar are generated by TAS software (Affymetrix). signal.bar contains log2(IP/Input) fold changes for each probe in a region 2*sliding window+1. pvalue.bar contains -10log10 (IP/Input) for each probe in a region 2*sliding window+1.
 
Submission date Aug 25, 2011
Last update date Nov 28, 2011
Contact name Thomas Montavon
Organization name EPFL
Street address SV 2843, Station 19
City Lausanne
ZIP/Postal code 1015
Country Switzerland
 
Platform ID GPL14183
Series (2)
GSE31658 Spatial organization of chromatin at the HoxD locus in developing limbs and brain
GSE31659 HoxD locus in the developing limb, digit, and brain

Supplementary file Size Download File type/resource
GSM786046_Input_1.CEL.gz 539.6 Kb (ftp)(http) CEL
GSM786046_Input_2.CEL.gz 545.9 Kb (ftp)(http) CEL
GSM786046_Limb_isIV_1+2_pvalue.bar.gz 451.3 Kb (ftp)(http) BAR
GSM786046_Limb_isIV_1+2_signal.bar.gz 291.0 Kb (ftp)(http) BAR
GSM786046_Limb_isIV_1.CEL.gz 520.1 Kb (ftp)(http) CEL
GSM786046_Limb_isIV_2.CEL.gz 544.1 Kb (ftp)(http) CEL
Processed data provided as supplementary file

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