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Sample GSM7585108 Query DataSets for GSM7585108
Status Public on Dec 27, 2023
Title IgG_1
Sample type SRA
 
Source name Lens
Organism Gallus gallus
Characteristics tissue: Lens
antibody: IgG
treatment: Embryonic day 4.5
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.
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.
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model HiSeq X Ten
 
Description Biosample A
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
 
Submission date Jul 10, 2023
Last update date Dec 27, 2023
Contact name Katia Del Rio-Tsonis
E-mail(s) delriok@miamioh.edu
Organization name Miami University
Department Biology
Street address 700 E High St
City Oxford
State/province OH
ZIP/Postal code 45056
Country USA
 
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.
Relations
BioSample SAMN36379442
SRA SRX20959235

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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