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| Status |
Public on Mar 28, 2024 |
| Title |
BJ1, Input |
| Sample type |
SRA |
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| Source name |
mouse embyronic stem cells
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| Organism |
Mus musculus |
| Characteristics |
tissue: mouse embyronic stem cells
cell line: BJ1 (C57BL/6J x JF1)
cell type: mouse embyronic stem cells
strain: C57BL/6J x JF1
chip antibody: none (Input)
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| Growth protocol |
ESGRO Complete PLUS Clonal Grade Medium (Merck)
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
BJ1 and JB1 ESCs for ChIP-seq experiment were fixed for 5 min in 2% formaldehyde at room temperature while tumbling. A 15 million cell pellet was sequentially lysed in the following 3 buffers: 5mL Buffer 1 (10% Glycerol, 50 mM Tris pH7.5, 140 mM NaCl, 1 mM EDTA, 0.5% NP40, 0.25% Triton X100, 0.33X protease inhibitors (Roche)) for 10 min on ice; 5mL Buffer 2 (10 mM Tris pH7.5, 0.2 M NaCl, 1 mM EDTA, 0.33X Protease inhibitors (Roche)) for 10 min on ice; 1mL Buffer 3 (100 mM NaCl, 10 mM Tris pH7.5, 1 mM EDTA, 0.25% SDS, 0.1% Sodium Deoxycholate, 1X Protease inhibitor (Roche)) for 10 min on ice. The chromatin was then sheared for 15 min using a Covaris S220 device (Peak power: 140W; Duty Factor: 20%; Cycle/Burst: 200), clarified by centrifugation, quantified and stored at -80°C. For CTCF ChIP-seq experiment, 10 µg sonicated chromatin was diluted in the following buffer (16 mM Tris pH8, 1 mM EDTA, 167 mM NaCl, 1.1% Triton X100, 0.01% SDS, 1X Protease inhibitor (Roche)) to a total volume of 1mL and was precipitated with 5 µg CTCF antibody (Millipore # 07-729) overnight. After recovering the antibody-bound chromatin with Salmon Sperm DNA/Protein A agarose beads (Millipore #16-157), beads were sequentially washed in Low Salt Wash buffer (20 mM Tris pH8.0, 150 mM NaCl, 2 mM EDTA, 1% Triton X100, 0.1% SDS), High Salt Wash buffer (20 mM Tris pH8.0, 500 mM NaCl, 2 mM EDTA, 1% Triton X100, 0.1% SDS), LiCl Wash buffer (10 mM Tris-HCl pH 7.5, 1 mM EDTA, 0.26 M LiCl, 1% NP-40, 1% Na Deoxycholate) and twice in TE buffer (10 mM Tris pH7.5, 1 mM EDTA). Chromatin was then eluted in 1% SDS, 0.1 mM NaHCO3 and decrosslinked over-night.
ChIP-seq libraries were prepared on purified DNA using the NEB Ultra II DNA library prep kit following manufacturer’s instructions
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
NextSeq 550 |
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| Data processing |
For Allele-specific analysis: Reads were aligned with bowtie2 on an mm10 N-masked genome in which SNPs differing between JF1 and C57BL/6J genomes were replaced by an N, the list of SNP being been obtained from the Mouse Genomes Project (release #1807). SNPsplit tool (Krurger et al. 2016; PMID: 27429743) was used to allocate reads to their respective allele and bedgraph for uniquely mapped and paired reads were generated using bedtools for each allele. Bedgraphs were scaled to a total of 10 million uniquely mapped and paired reads on both JF1 and C57BL/6J allele. A bedgraph has also been generated with unassigned reads (and scaled to a total of 10 million uniquely mapped and paired reads)
For non-allelic analysis (total): Reads were aligned with bowtie2 on mm10 genomes. The corresponding bedgraph was generated using bedtools (scaled to a total of 10 million uniquely mapped and paired reads).
Assembly: mm10
Supplementary files format and content: Enrichment score in bedgraph format (mm10)
Supplementary files format and content: A bedgraph for each allele (JF1 and Bl6). A bedgraph for unassigned reads (not overlapping known SNPs). A bedgraph for total reads (non-allelic)
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| Submission date |
Jun 29, 2022 |
| Last update date |
Mar 28, 2024 |
| Contact name |
Benoit Moindrot |
| E-mail(s) |
benoit.moindrot@i2bc.paris-saclay.fr
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| Organization name |
Institute for Integrative Biology of the Cell
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| Street address |
1, avenue de la terrasse
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| City |
Gif-sur-Yvette |
| ZIP/Postal code |
91198 |
| Country |
France |
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| Platform ID |
GPL21626 |
| Series (1) |
| GSE207166 |
The long non-coding RNA Meg3 mediates imprinted gene expression during stem cell differentiation |
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| Relations |
| BioSample |
SAMN29417870 |
| SRA |
SRX15940605 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6280761_BJ_Input_masked_genome1.Bl6_Coverage.txt.gz |
321.8 Mb |
(ftp)(http) |
TXT |
| GSM6280761_BJ_Input_masked_genome2.JF1_Coverage.txt.gz |
297.6 Mb |
(ftp)(http) |
TXT |
| GSM6280761_BJ_Input_masked_unassigned_Coverage.txt.gz |
1.1 Gb |
(ftp)(http) |
TXT |
| GSM6280761_BJ_Input_total_Coverage.txt.gz |
1.7 Gb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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