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| Status |
Public on Mar 13, 2023 |
| Title |
DLD-1 cln22 H3K4me3 biol rep 2 |
| Sample type |
SRA |
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| Source name |
DLD-1 Cells
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| Organism |
Homo sapiens |
| Characteristics |
cell line: DLD-1 Cells
cut&run antibody: H3K4me3 - EpiCypher (13-0041)
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| Treatment protocol |
n/a (clones and parental cells were obtained from the authors of Ly et al., Nat. Genet. 2019)
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| Growth protocol |
DLD-1 cells were grown in RPMI supplemented with 10% FBS and 1X Pen-Strep in 100 mm dish (Corning Corning™ 100 mm TC-Treated Culture Dishes)
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
CUTANA CUT&RUN, Illumina sequencing, and data analysis. CUTANA(R) CUT&RUN was performed on an automated protocol (autoCUT&RUN) derived from those previously described [1-3]. In brief, for each CUT&RUN reaction 500K cells [5 million cells/mL prepared in 20 mM HEPES, pH 7.5, 150 mM NaCl, 0.5 mM Spermidine, 1 mM Trichostatin A, 1X EDTA/EGTA-free Complete Protease Inhibitor] were dispensed to individual wells of a 96-well plate, immobilized onto Concanavalin-A beads (Con-A; EpiCypher #21-1401), and incubated overnight (4°C) with 0.5 µg of antibody (IgG, H3K4me3, H3K27me3, H3K27ac) (all antibodies validated to histone post-translation modification (PTM)-defined SNAP-ChIP nucleosome standards as previously [4]). pAG-MNase (EpiCypher #15-1016) was added / activated (2 hours @ 4°C) and CUT&RUN enriched DNA purified using Serapure beads after mixing at 2:1 (Bead:DNA) ratio. Recovered DNA was quantified using PicoGreen and reactions were normalized to 5ng DNA (or entirety of the reaction if <5ng DNA was recovered) before preparing sequencing libraries (CUTANA CUT&RUN Library Prep kit;EpiCypher #14-1001). All autoCUT&RUN steps were optimized / performed on Tecan Freedom EVO robotics platforms with gentle rocking for incubation steps and magnetic capture for media exchange / washing steps. • [1] Marunde et al (2022) bioRxiv [doi.org/10.1101/2022.02.21.481373] • [2] Yusufova et al (2021) Nature 589:299 [PMID: 33299181] • [3] Skene et al (2018) Nat Protoc 13:1006 • [4] Shah et al (2018) Molecular Cell 72:172 [PMID: 30244833]
Library was prepared according to Epicypher CUT&RUN Kit library prep manual https://www.epicypher.com/content/documents/manuals/14-1001-2-cut-and-run-library-prep-kit-manual.pdf
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
NextSeq 2000 |
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| Data processing |
Alignment performed using Bowtie2 v. 2.2.5 Blacklist regions from Encode. Cite: Amemiya, H.M., Kundaje, A. & Boyle, A.P. The ENCODE Blacklist: Identification of Problematic Regions of the Genome. Sci Rep 9, 9354 (2019). https://doi.org/10.1038/s41598-019-45839-z
Bigwigs generated using Deeptools v. 3.5.1
Peak calling performed with MACS2 v. 2.2.7.1
Peaks annotated with HOMER
Assembly: hg38
Supplementary files format and content: bigwig
Library strategy: CUT&RUN
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| Submission date |
Sep 23, 2022 |
| Last update date |
Mar 13, 2023 |
| Contact name |
Albert S Agustinus |
| Organization name |
Memorial Sloan Kettering Cancer Center
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| Street address |
408 East 69th Street, Room 419-G
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| City |
New York |
| State/province |
New York |
| ZIP/Postal code |
10021 |
| Country |
USA |
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| Platform ID |
GPL30173 |
| Series (2) |
| GSE186589 |
Epigenetic dysregulation from chromosomal transit in micronuclei |
| GSE214012 |
Epigenetic dysregulation from chromosomal transit in micronuclei [DLD-1 CUT&RUN] |
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| Relations |
| BioSample |
SAMN30983542 |
| SRA |
SRX17681431 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6597833_CB100_H3K4me3_N_DLD-1_cln22_500K_KMET_S100_uniq.b20s100dt.bw |
80.8 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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