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| Status |
Public on Dec 15, 2016 |
| Title |
mouse_SOX9_ChIPseq_1 |
| Sample type |
SRA |
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| Source name |
fetal testis
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| Organism |
Mus musculus |
| Characteristics |
tissue: testis
strain: swiss OF1
developmental stage: E13.5
antibody: rabbit anti SOX9 homemade (ref Gasca et al, PNAS 2002, vol 99, 11199-11204)
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| Growth protocol |
Animal care and handling were conformed to the “Réseau des Animaleries de Montpellier” (RAM). For bovine fetus production and collection, experiments reported in this work were in agreement with the ethical guidelines of the French National Institute for Agricultural Research (INRA). Fetuses were produced by artificial insemination of Holstein females with semen of Holstein males (day 0), then collected at 90 days post-fertilization at the INRA slaughterhouse (France). The protocol (N°: 12/046) was approved by the local ethical committee (COMETHEA) and Eric Pailhoux is the recipient of an official authorization for animal experimentation (N°: B91-649).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Upon collection, gonads were crushed in liquid nitrogen using a mortar. Tissue powders were immediately fixed with PBS containing 2mM of Disuccinimidyl glutarate (DSG: PIERCE ref 20693) initially solubilized in DMSO as described previously (Nowak et al. 2005). Samples were then incubated for 30 minutes on a rotating wheel at room temperature. After three washes with PBS, samples were subsequently fixed with PBS 1% formaldehyde for 30 minutes at room temperature
Libraries were prepared according to Illumina's instructions. 10-300ng DNA is combined with End Repair Mix, and incubate at 20°C for 30 min. Purify the end-repaired DNA with QIAquick PCR Purification Kit(Qiagen),then add A-Tailing Mix, incubate at 37°C for 30 min. Combine the purified Adenylate 3'Ends DNA, Adapter and Ligation Mix, incubate the ligation reaction at 20°C for 15 min. Purify the Adapter-ligated DNA with the QIAquick PCR Purification Kit. Several rounds of PCR amplification with PCR Primer Cocktail and PCR Master Mix are performed to enrich the Adapter-ligated DNA fragments. Then the PCR products are selected by running a 2% agarose gel to recover the target fragments. Purify the gel with QIAquick Gel Extraction kit (QIAGEN).
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
processed data file: Mouse_SOX9_ChIPseq.bed
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| Data processing |
The base calling software name version and parameters are OLB-1.9.4/bin/setupBclToQseq.py --input-dir Data/Intensities/BaseCalls --output-dir --use-bases-mask Y*,Y*,Y*
ChIP-seq reads were aligned to the mouse mm9 or bovine Bostau6 (UMD3.1) genome assembly using SOAP2.21
For peaks calling, the number of reads being twice in the inputs than in the ChIP-seq experiments, we generated from the inputs a random sampling of 20 million reads each, using a homemade script which respects the proportion of reads per chromosome of the initial input. With this method we generated ten input samples for each ChIP-seq analysis that were used for ten rounds of peaks calling with the MACS software (1.4) (Zhang et al. 2008)( bw=250, mfold=10 et p value=1e-5). Peaks common to the ten operations with FDR<0.05 were kept for further analysis. Script is available upon request
Genome_build: mm9 or BosTau6
Supplementary_files_format_and_content: bed with peaks coordinate in mm9 for mouse and BosTau6 for bovine
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| Submission date |
May 16, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Francis Poulat |
| E-mail(s) |
francis.poulat@igh.cnrs.fr
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| Phone |
(+33) 4 3435 9940
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| Organization name |
IGH CNRS-UM UMR9002
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| Department |
Genetic and Development
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| Street address |
141 rue de la Cardonille
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| City |
Montpellier |
| ZIP/Postal code |
34396 |
| Country |
France |
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| Platform ID |
GPL13112 |
| Series (2) |
| GSE81488 |
In fetal testis, SOX9 acts on transcription and splicing of its targets genes through binding to genomic regions with conserved signatures [ChIP-seq] |
| GSE81490 |
In fetal testis, SOX9 acts on transcription and splicing of its targets genes through binding to genomic regions with conserved signatures |
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| Relations |
| BioSample |
SAMN05003705 |
| SRA |
SRX1769357 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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