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| Status |
Public on Feb 17, 2023 |
| Title |
CUTRUN_H3K27ac_67-Pupa_bioRep-1 |
| Sample type |
SRA |
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| Source name |
Pupal wings
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| Organism |
Drosophila melanogaster |
| Characteristics |
tissue: Pupal wings
developmental stage: 67% of pupal development
Sex: Male
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| Growth protocol |
D. melanogaster line D2, a transgenic line used in previous study (GSE142176), was maintained on standard cornmeal medium at 25ºC with a 12:12 day-night light cycle.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
35-45 wings per biological replicate at 67% pupal development from D. melanogaster line D2 were transferred into a 5 ml Eppendorf tube with cold 1x PBS and 6 mM MgCl2, rocked at 4°C with 1x cold lysis buffer for 5 min and then lightly crosslinked (1 min) with the crosslink buffer. After light crosslink, the wings were immediately quenched with glycine, and then transferred into a glass well with 50 μl cold XL WB buffer and cut coarsely (3 pieces per wing). Homogenization was performed with a 2 ml Dounce using pestle A for 12 strokes and then pestle B for 30 strokes. The homogenate was incubated on ice for 40-50 min, and then centrifuged at 1000 g for 10 min at 4°C with an addition of 500 μl of XL WB. The nuclei pellet was resuspended with 100 μl XL WB and processed for CUT&RUN. The CUT&RUN steps were performed following the instructions from EpiCypher, including bead activation, binding cells to activated beads, binding of antibodies, binding of pAG-MNase, targeted chromatin digestion and release, and reverse crosslinking. 1 μl of antibody against H3K27ac (Active Motif #39034) was used. DNA was then purified with Qiagen MinElute kit and subsequently processed for library preparation.
Libraries were prepared using NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® and NEBNext® Multiplex Oligos for Illumina® (Dual Index Primers Set 2) following a protocol published on protocols.io (DOI: dx.doi.org/10.17504/protocols.io.wvgfe3w).
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 1500 |
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| Data processing |
library strategy: CUT&RUN
Libraries were demultiplexed and trimmed, and then aligned to the reference genome UCSC dm6 using Bowtie2 with following parameters: --end-to-end --very-sensitive --no-mixed --no-discordant -q --phred33 -I 10 -X 700.
The aligned reads were filtered and cleaned as previously described (GSE142176).
Peak calling was performed by MACS2 with following settings: -f BAMPE --keep-dup all -q 0.01 -g 1.2e+8 -B --SPMR.
BigWig files were converted from normalized bedGraph files generated by MACS2 above.
Assembly: dm6
Supplementary files format and content: BigWig files for normalized signal track,narrowpeak files for peaks called by MACS2.
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| Submission date |
Jan 12, 2023 |
| Last update date |
Feb 17, 2023 |
| Contact name |
Nicolas Gompel |
| Organization name |
Ludwig Maximilian University of Munich
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| Department |
Faculty of Biology
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| Street address |
Grosshaderner Str. 2
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| City |
Planegg-Martinsried |
| ZIP/Postal code |
82152 |
| Country |
Germany |
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| Platform ID |
GPL19951 |
| Series (2) |
| GSE222715 |
Increased chromatin accessibility promotes the evolution of a transcriptional silencer in Drosophila [CUT&RUN] |
| GSE222717 |
Increased chromatin accessibility promotes the evolution of a transcriptional silencer in Drosophila. |
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| Relations |
| BioSample |
SAMN32691651 |
| SRA |
SRX19014211 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6929278_CUTRUN_H3K27ac_67-Pupa_bioRep-1.bw |
75.0 Mb |
(ftp)(http) |
BW |
| GSM6929278_CUTRUN_H3K27ac_67-Pupa_bioRep-1.narrowpeak.gz |
368.1 Kb |
(ftp)(http) |
NARROWPEAK |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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