|
| Status |
Public on Feb 17, 2021 |
| Title |
NEI002-I138fs-Patient-D200-1 |
| Sample type |
SRA |
| |
|
| Source name |
Retinal Organoid
|
| Organism |
Homo sapiens |
| Characteristics |
age: d200
genotype: CRX-I138fs
|
| Treatment protocol |
All samples were cultured in identical conditions.
|
| Growth protocol |
Retinal organoids were derived using a combination of 2D and 3D culture. Embryoid bodies were formed from iPSCs by EDTA treatment, cultured for 7 days in neural induction medium followed by plating on Matrigel. Between 3-4 weeks neural retina domains were dissected from adherent cultures forming organoids, which were subsequently maintained as floating cultures in ultra-low attachment 96-well plate format until collection.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Organoids were snap frozen on dry ice at the indicated day of differentiation. Frozen samples were thawed on ice and RNA extracted using Qiagen Rneasy Mini Kit according to the manufacturer's manual. RNA quality was determined using Agilent Bioanalyzer, all samples had RIN >7.
RNA was isolated from 3 frozen organoids per sample using QIAGEN RNeasy Kit according to manufacturer’s manual. RNA quality was assessed using Bioanalyzer (Agilent Technologies). Samples with RIN >7 were subsequently used for library generation using TruSeq Library Prepartation Kit (Illumina Inc.).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
CRX disease modeling in retinal organoid (I138fs mutation)
I138fs_raw_counts.txt
I138fs_norm_counts_5CPM.txt
|
| Data processing |
Fastq files were generated from reads passing chastity filter.
Transcript-level quantification was performed with kallisto v0.45.0 against Ensembl release 94
Transcript counts were summarized to the gene level using tximport using the lengthscaledTPM option (raw_counts.txt files)
Genes with <5cpm were removed and remaining genes were TMM-normalized and converted to counts per million (CPM) (norm_counts_5CPM.txt files)
Genome_build: GRCh38.p12
Supplementary_files_format_and_content: raw count txt file, abundance measurements txt file
|
| |
|
| Submission date |
Jun 22, 2020 |
| Last update date |
Nov 09, 2023 |
| Contact name |
Zachary A Batz |
| Organization name |
NIH
|
| Department |
NEI
|
| Lab |
NNRL
|
| Street address |
6 Center Dr Bldg 6, Rm 303
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (2) |
| GSE152939 |
Gene therapy of dominant CRX-Leber congenital amaurosis using patient retinal organoids I |
| GSE153101 |
Gene therapy of dominant CRX-Leber congenital amaurosis using patient retinal organoids |
|
| Relations |
| BioSample |
SAMN15339938 |
| SRA |
SRX8594511 |
