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| Status |
Public on Jun 22, 2022 |
| Title |
H3K9/14ac_HCwater_rep1 |
| Sample type |
SRA |
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| Source name |
hippocampus
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| Organism |
Mus musculus |
| Characteristics |
chip antibody: H3K9/14ac (Diagenode, C15410200)
agent: water
strain: C57Bl6/J
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| Treatment protocol |
Two-three month-old mice were randomly assigned to the two following experimental groups: water (control group) and caffeine. The caffeine solution was kept in dark bottles thus protected from light and changed weekly. Treatment started at eight to nine weeks of age and lasted for two weeks. The chronic caffeine treatment in mice has been set in order to mimic the usual dose range of caffeine consumption in Humans. The selected caffeine dose of 0.3 g/L p.o., administered through drinking water at 0.3 g/L, has been previously shown to provide a significant benefit in neurodegenerative contexts (Arendash et al., 2006; Arendash et al., 2009; Laurent et al., 2014).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Freshly dissected tissue was chopped by a razor blade and rapidly incubated in 1.5 mL PBS containing 1% formaldehyde for 10 minutes at RT. To stop fixation, glycine was added (0.125 M final concentration). Dorsal hippocampi from 4 mice were pooled per sample. Tissue samples were then processed as described in Chatterjee et al. 2018, and sonicated using the Diagenode Bioruptor (30 s ON-30 s OFF at High Power x 35 cycles). Sonicated chromatin was centrifuged 10 min at 14000xg, the supernatant collected and diluted 1:10 in ChIP dilution buffer (0.01% SDS, 1.1% Triton X-100, 1.2 mM EDTA, 16.7 mM Tris-Cl, pH 8.1, 167 mM NaCl). A fraction of the supernatant (50 mL – 10%) from each sample was saved before immune-precipitation for ‘total input chromatin’. Supernatants were incubated overnight (4°C) with 1/1000 primary antibody H3K9/14ac (Diagenode C15410200) and H3K27ac (ab4729), followed by protein A Dynabeads (Invitrogen) for 2 hours at RT. After several washes (low salt, high salt, LiCl and TE buffers), the resulting DNA-protein complexes were eluted in 300 mL elution buffer (1% SDS, 0.1 M NaHCO3). The crosslinking was reversed (overnight at 65°C) and the DNA was purified with RNAse (30 min, 37°C), proteinase K (2 hours at 45°C). DNA from the immunoprecipitated and input samples was isolated using Diagenode MicroChIP DiaPure columns with 20 mL nuclease-free milliQ water in low binding tubes.
ChIP samples were purified using Agencourt AMPure XP beads (Beckman Coulter) and quantified with the Qubit (Invitrogen). ChIP-seq libraries were prepared from 10ng of double-stranded purified DNA using the MicroPlex Library Preparation kit v2 (C05010014, Diagenode s.a., Seraing, Belgium), according to manufacturer's instructions. In the first step, the DNA was repaired and yielded molecules with blunt ends. In the next step, stem-loop adaptors with blocked 5 prime ends were ligated to the 5 prime end of the genomic DNA, leaving a nick at the 3 prime end. The adaptors cannot ligate to each other and do not have single-strand tails, avoiding non-specific background. In the final step, the 3 prime ends of the genomic DNA were extended to complete library synthesis and Illumina compatible indexes were added through a PCR amplification (4 + 7 cycles). Amplified libraries were purified and size-selected using Agencourt AMPure XP beads (Beckman Coulter) to remove unincorporated primers and other reagents.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 4000 |
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| Description |
H3K9K14ac water
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| Data processing |
Image analysis and base calling were performed using RTA 2.7.3 and bcl2fastq 2.17.1.14.
Sequence reads were mapped to reference genome mm10 using Bowtie 1.0.0 with the following parameters -m 1 --strata --best -y -S -l 40 -p 2.
Genome_build: mm10
Supplementary_files_format_and_content: Wig files were generated using with an in-house script (Variable step, span=25, reads were elongated to 200b).
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| Submission date |
Feb 19, 2021 |
| Last update date |
Oct 27, 2023 |
| Contact name |
David Blum |
| E-mail(s) |
[email protected]
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| Organization name |
Inserm
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| Department |
Lille Neurosience & Cognition
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| Lab |
Alzheimer & Tauopathies
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| Street address |
1 place de Verdun
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| City |
Lille |
| State/province |
Cedex |
| ZIP/Postal code |
59045 |
| Country |
France |
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| Platform ID |
GPL21103 |
| Series (2) |
| GSE167122 |
Genomewide effects of regular caffeine intake on hippocampal metabolism and learning-dependent transcription [ChIP-Seq] |
| GSE167123 |
Genomewide effects of regular caffeine intake on hippocampal metabolism and learning-dependent transcription |
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| Relations |
| BioSample |
SAMN18006319 |
| SRA |
SRX10134699 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5097093_wigs_for_DDBM86.wig.gz |
222.5 Mb |
(ftp)(http) |
WIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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