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| Status |
Public on Oct 12, 2022 |
| Title |
ATAC NT Rep 1 |
| Sample type |
SRA |
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| Source name |
Schneider 2 (S2) cells
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| Organism |
Drosophila melanogaster |
| Characteristics |
strain: Oregon R
developmental stage: Late embryonic stage
antibody: N/A
spike-in: Drosophila virilis
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| Treatment protocol |
Experiments were conducted on cells at 80-90% confluency. Cells were resuspended in FBS-free media and incubated with 30 μM of C646 for 1 hr or 4 hr.
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| Growth protocol |
S2 cells were cultured at 25°C in Schneider's Drosophila medium (Gibco) supplemented with 10% fetal bovine serum (FBS) (Gibco) and 1% Penicillin-Streptomycin (Gibco).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Crude nuclear extracts were isolated from 100,000 untreated and C646-treated S2 cells by resuspension in ATAC lysis buffer (10 mM Tris pH 7.4, 10 mM NaCl, 3 mM MgCl2 and 0.1& IGEPAL CA-630) and centrifugation at 700 g for 10 min. 10,000 spike-in nuclei from 2-4 h D. virilis embryos (dechorionated and crude nuclear extracts prepared by manual homogenization with a pestle in ATAC lysis buffer) were added to the S2 nuclear pellets which were resuspended in 22.5 μl ATAC lysis buffer, 2.5 μl Tn5 (Tagment DNA Enzyme 1(TDE1) (Illumina)) and 25 μl Tagment DNA Buffer (Illumina). Tagmentation was performed at 37°C for 30 min on a thermomixer at 1,000 rpm. The transposition reaction was stopped by addition of 1% SDS and DNA purified by the addition of Agencourt AMPure XP beads (Beckman Coulter A63881) at a 2:1 ratio of beads to sample. Purification was performed following the manufacturer’s instructions.
Tagmented DNA was PCR amplified using 1x Phusion® High-Fidelity PCR Master Mix with GC Buffer (NEB), 1.25 μM PCR Primer Cocktail (Illumina) and 1.25 μM Nextera PCR primers i5 and i7 (Nextera® Index Kit (Illumina)). PCR amplification conditions were: 72°C for 5 min, followed by 10 cycles of 98°C 10 seconds, 65°C 1 min and 15 seconds, then 72°C for 1 min. Amplified libraries were purified with Agencourt AMPure XP beads (Beckman Coulter A63881) using a 1:5:1 ratio of bead to sample volume. Libraries were sequenced paired-end (2x 37 bp) on the Illumina NextSeq 550 platform at the BEA core facility, Stockholm.
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| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
NextSeq 550 |
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| Description |
ATAC NT Rep 1
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| Data processing |
ATAC-seq reads from each sequenced library were mapped to the Drosophila melanogaster (dm6) genome assembly using Bowtie2 with the default parameters. Duplicated and unmapped reads were removed using Picard. To obtain the spike-in reads, we mapped the reads to the Drosophila virilis (dvir_caf1) genome assembly and the D. melanogaster reads were normalized based on the ratio of D. virilis reads. From the spike-in adjusted data, RPKM normalized ATAC-seq coverage tracks were generated using the deepTools. Bigwig files of the mean RPKM were produced using the deepTools comparebigwig tool.
Genome_build: dm6
Supplementary_files_format_and_content: bigwig for coverage
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| Submission date |
Oct 27, 2021 |
| Last update date |
Oct 12, 2022 |
| Contact name |
Mattias Mannervik |
| E-mail(s) |
mattias.mannervik@su.se
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| Organization name |
Stockholm University
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| Department |
Molecular Biosciences, the Wenner-Gren Institute
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| Lab |
Mattias Mannervik
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| Street address |
Arrheniuslaboratories E3
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| City |
Stockholm |
| ZIP/Postal code |
10691 |
| Country |
Sweden |
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| Platform ID |
GPL22106 |
| Series (2) |
| GSE186720 |
p300/CBP sustains Polycomb silencing by non-enzymatic functions (ATAC-seq on inhibitor-treated Drosophila S2 cells) |
| GSE186723 |
p300/CBP sustains Polycomb silencing by non-enzymatic functions |
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| Relations |
| BioSample |
SAMN22608643 |
| SRA |
SRX12801964 |
| 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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