|
| Status |
Public on Dec 31, 2020 |
| Title |
FD19 |
| Sample type |
SRA |
| |
|
| Source name |
After form deprivation myopia (FDM) for 7 days, occluders removed to allow 6 hrs of normal light and FDM recovery (FDMR)
|
| Organism |
Gallus gallus |
| Characteristics |
age: Post-hatch day 20
time: 7 days (post-occlusion)
lens type: FDMR (6hrs)
tissue: Retina, RPE, choroid
|
| Treatment protocol |
Male chicks (Leghorn/New Hampshire) were raised from post-hatch days 0-5 under a 12-hr day/night light cycle in groups of <25. In the middle of the light cycle on day 5, chicks were randomly assigned to a lens condition (FD or No Lens) and occluders attached to Velcro were fixed to the periocular feathers of the right eye. Following a further 7 days, tissue was collected from no lens and FDM chicks. At the same time, occluders were removed from FDMR chicks. Tissue was collected from these FDMR chicks following 6hrs or 24hrs recovery. Description of tissue collection: Chicks were anaesthetized and right eye refraction and axial dimensions determined by retinoscopy and A-Scan ultrasonography. Chicks were euthanized and their right eyes were enucleated. The retina/RPE/choroid was immediately collected from the posterior eyecup and frozen in liquid nitrogen before being transferred to -80°C.
|
| Growth protocol |
N/A
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from the retina/RPE/choroid using the miRNeasy Mini Kit (Qiagen) including DNase digestion. RNA quality and quantity was assessed on the 2100 Bioanalyzer. All samples had an RNA integrity number (RIN) of >8. RNA quantity was also assessed on the Qubit 2.0 Fluorometer.
Using an average of concentration measures obtained from Qubit and Bioanalyzer assays, 2.5μg of RNA from each sample was used for library preparation. Libraries were prepared using the TruSeq Stranded mRNA LS kit (Illumina) with dual indexing according to the manufacturer’s instructions. The generated libraries were assessed on the 2100 Bioanalyzer to ensure an average size distribution of approximately 280bps, then quantified on the Qubit 2.0 Fluorometer and by qPCR. Libraries were normalised to 10nM in Tris-HCl, pooled, and prepared for cluster generation on the Illumina cBot using the TruSeq SR Cluster Kit V3-cBot (Illumina) with denatured template DNA diluted to 7pM. The flow cell and sequencing reagents (TruSeq SBS Kit V3; Illumina) were loaded on the Illumina HiSeq 1500 and a dual-index, single-end, 100bp sequencing run performed.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 1500 |
| |
|
| Description |
After initiation of form deprivation myopia (FDM) for 7 days, occluders were removed to allow 6 hrs of normal light and FDM recovery (FDMR)
|
| Data processing |
Read quality was assessed using FastQC
Adapter and low quality (Q-score <10) sequences removed using CutAdapt and Trimmomatic
Reads were mapped to the chick genome using Tophat2 and Bowtie2
The number of reads uniquely mapping to each gene was counted using existing gene models with HTSeq
Genome_build: GalGal4
Supplementary_files_format_and_content: The txt file contains processed data for all samples (raw counts)
|
| |
|
| Submission date |
Apr 15, 2016 |
| Last update date |
Dec 31, 2020 |
| Contact name |
Loretta Giummarra |
| E-mail(s) |
L.Giummarra@latrobe.edu.au
|
| Organization name |
La Trobe University
|
| Department |
Psychology & Counselling
|
| Street address |
c/o Psychological Science
|
| City |
Melbourne |
| State/province |
Victoria |
| ZIP/Postal code |
3086 |
| Country |
Australia |
| |
|
| Platform ID |
GPL21476 |
| Series (1) |
| GSE80327 |
RNA-seq during occlusion myopia induction and recovery |
|
| Relations |
| BioSample |
SAMN04859602 |
| SRA |
SRX1707260 |
