|
| Status |
Public on May 11, 2022 |
| Title |
ATAC-Seq E4 RPE 2 |
| Sample type |
SRA |
| |
|
| Source name |
Retinal pigment epithelium
|
| Organism |
Gallus gallus |
| Characteristics |
treatment: Development
Stage: E4
|
| Treatment protocol |
For treated conditions, surgical retinectomy was performed and FGF2-containing heparain beads were delivered to the eye cup at the time of injury.
|
| Growth protocol |
White Leghorn chicken eggs obtained from Michigan State University were used. Eggs were incubated in a humidified, rotating incubator at 38 °C before performing experiments at Hamburger Hamilton stage 24 (E4) or 26 (E5).
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
RPE was isolated in cold PBS and two isolated sheets of RPE were collected per biological sample and placed on ice. RPE cells were lysed in cold ATAC-Seq Kit lysis buffer (Active Motif cat. 53150) and nuclei were counted with a hemocytometer.
Approximately 100,000 RPE nuclei from 2 embryos were aliquoted for Tn5 tagmentation per sample and ATAC-seq library preparation was performed using ATAC-Seq Kit (Active Motif cat. 53150) per manufacturer’s instructions.
|
| |
|
| Library strategy |
ATAC-seq |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
E4_RPE2
|
| Data processing |
Reads were trimmed using cut adapt and trim galore with the trimming parameters --clip_R1 16 --clip_R2 18 --three_prime_clip_R1 6 --three_prime_clip_R2 4.
Alignment of raw reads to chicken genome GRCg6a was performed with Bowtie2 (Langmead and Salzberg, 2012) using the parameters --very-sensitive -k 5 -p 40.
Aligned reads were deduplicated using Picard tools (Broad Institute, Picard Toolkit).
Peaks were called for samples individually with HMMRATAC (Tarbell and Liu, 2019). HMMRATAC defined open regions were unified across all samples into a consensus peak file using BEDtools merge -d 10 -I (Quinlan and Hall, 2010).
Peaks aligning to the mitochondrial chromosome were discarded, and peak counts were determined using featureCounts with parameters -F SAF -s 0 -T 10 (Liao et al., 2014).
Differential expression testing of peak regions and peak summits was performed with DESeq2.
Genome_build: GRCg6a
Supplementary_files_format_and_content: Normalized peak count matrix from DESeq2.
|
| |
|
| Submission date |
Mar 04, 2022 |
| Last update date |
May 11, 2022 |
| 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 |
GPL23499 |
| Series (2) |
| GSE197935 |
Chromatin accessibility (ATAC-seq) in the developing chicken retinal pigment epithelium |
| GSE197938 |
OTX2 regulatory network and maturation-associated gene programs are inherent barriers to RPE neural competency |
|
| Relations |
| BioSample |
SAMN26440711 |
| SRA |
SRX14371932 |
