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| Status |
Public on Dec 27, 2023 |
| Title |
H3K4me3_2 |
| Sample type |
SRA |
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| Source name |
Lens
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| Organism |
Gallus gallus |
| Characteristics |
tissue: Lens
antibody: H3K4me3
treatment: Embryonic day 4.5
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| Growth protocol |
Chicken embryos were obtained from Michigan State University and lenses were dissected from embryonic day 4.5 chicken embryos. Dissected lenses were collected on ice and pooled into 3 biological replicates consisting of approximately 44 lenses each.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
In brief, nuclei extraction was carried out via an optimized variation of the demonstrated protocols described in 10X Genomics Protocol CG000124 Rev E and 10X Genomics Protocol CG000366 Rev D. Cell lysis was performed by triturating sample and incubating in a cold buffer comprised of lysis was performed via trituration and 30 second incubation in a buffer containing 10 mM Tris-HCl, 10 mM NaCl, 3 mM MgCl2, 0.005% Tween-20, 0.001% Surfact-Amps NP-40, 0.0005% digitonin (Thermo Fisher Scientific, BN2006), 1% fraction V bovine serum albumin, 1X proteinase inhibitor (MilliporeSigma, 11873580001), and 1.5 mM spermidine. Nuclei were washed twice by resuspending in cold buffer containing 10 mM Tris-HCl, 10 mM NaCl, 3 mM MgCl2, 1% fraction V bovine serum albumin, 0.1% Tween-20, 1X proteinase inhibitor (MilliporeSigma, 11873580001), and 1.5 mM spermidine. Nuclei were passed twice through 20 µm cell strainers (pluriSelect, 43-10020-40) during isolation to filter debris and cell aggregates.
Nuclei across all samples were diluted to 1000 nuclei / µl and loaded at 100000 nuclei per reaction for CUT&RUN-seq using the CUTANA ChIC / CUT&RUN Kit Version 3 (EpiCypher, 14-1048) and CUTANA CUT&RUN Library Prep Kit (EpiCypher, 14-1002). The protocol was carried out according to manufacturer protocol, with slight modification. . Antibodies were delivered at the following concentrations: EZH2 (Cell Signaling, 5246) 1:100; SUZ12 (Cell Signaling, 3737) 1:100; H3K27me3 (Cell Signaling, 9733S) 1:50; H3K4me3 (EpiCypher, 13-0041k) 1:50; IgG (EpiCypher, 13-0042k) 1:50; JARID2 (Cell Signaling, 13594) 1:50; AEBP2 (Cell Signaling, 14129) 1:25.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
HiSeq X Ten |
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| Description |
Biosample B
H3K4me3_peaks.narrowPeak
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| Data processing |
library strategy: CUT&RUN-seq
Raw reads were quality and adapter trimmed using Trim Galore! (v0.6.4_dev) with parameters –phred33 –paired –length 36 -e 0.1 -q 5 –stringency 1
Trimmed reads were aligned against chicken genome GRCg7b with Bowtie 2 (v2.4.1) using parameters -X 2000 --local --no-mixed --no-discordant --no-dovetail
Peaks were called using MACS2 (v2.2.7.1) and parameters -f BAMPE -g 1070887886 --keep-dup all -q 0.05, providing IgG alignment files as background
Assembly: GRCg7b
Supplementary files format and content: Processed files contain narrow peak files generated using MACS2
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| Submission date |
Jul 10, 2023 |
| Last update date |
Dec 27, 2023 |
| Contact name |
Katia Del Rio-Tsonis |
| E-mail(s) |
delriok@miamioh.edu
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| Organization name |
Miami University
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| Department |
Biology
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| Street address |
700 E High St
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| City |
Oxford |
| State/province |
OH |
| ZIP/Postal code |
45056 |
| Country |
USA |
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| Platform ID |
GPL24517 |
| Series (2) |
| GSE236904 |
Integrated single-cell multiomics uncovers foundational regulatory mechanisms of lens development and pathology [CUT&RUN] |
| GSE236905 |
Integrated single-cell multiomics uncovers foundational regulatory mechanisms of lens development and pathology. |
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| Relations |
| BioSample |
SAMN36379439 |
| SRA |
SRX20959238 |
| 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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