|
| Status |
Public on Aug 22, 2020 |
| Title |
NSC-3 |
| Sample type |
SRA |
| |
|
| Source name |
neural stem cell (NSC)
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: NSCs diffrentiated from validated iPSCs
|
| Growth protocol |
iPSCs maintained in feeder-free conditions were split as cell clumps in a Geltrex coated six well plate at a density reaching 15-25% confluence after 24 hours and maintained in mTeSR-1 medium (Stem Cell Technologies) supplemented with 10 µM ROCK inhibitor Y27632 (ATCC® ACS3030™). Approximately 24 hours after splitting, culture medium was switched to Gibco PSC Neural Induction Medium (Life Technologies). Medium was changed every other day. On day 7 NSCs were dissociated with Stem Pro Accutase (Life Technologies) and plated on Geltrex coated plate at a density of 1 x 105 cells/cm2 in an NSC expansion medium containing 50% Neurobasal medium, 50% Advanced DMEM/F12, and 1X neural induction supplement (all from Life Technologies). The NSC expansion medium was supplemented with 5 µM ROCK inhibitor Y27632 (ATCC® ACS3030™) for first 24 hours. NSC expansion medium was changed every other day until NSCs reached confluence. The six generated NSC lines from passage one, were used for characterization and total RNA extraction.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA from snape frozen cell pellets (~3-5 x 106 cells) from six iPSC differentiated passage one NSCs was extracted using commercially available Rneasy Mini Kit (Qiagen) and the manufacturer protocol. RNA quality and quantity were assessed using a NanoDrop 2000 Spectrophotometer (Thermo Scientific) and an Agilent 2200 TapeStation system (Agilent Technologies).
1µg of total RNA per sample was used to prepare mRNA sequencing libraries. The sequencing libraries were prepared using Illumina TruSeq RNA sample preparation kit v2 and following manufacturer protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
NSC-mRNA.txt
|
| Data processing |
Raw fastq sequence files were generated and de-multiplexed using the Illumina CASAVA v1.8 pipeline.
After pre-alignment QCs, sequences were aligned to human genome assembly GRCh38 (hg38) and mapped to RefSeq transcripts using Strand NGS software v3.4 (Strand Genomics Inc.).
The aligned reads were then filtered based on read quality metrics and only perfectly aligned sequencing reads were retained and quantified using NGS software v3.4 (Strand Genomics Inc.).
The gene expression values as raw counts of sequencing reads were obtained and log2 transformation and ‘DESeq’ normalization was applied.
Genome_build: hg38
Supplementary_files_format_and_content: Tab-delimited text file of raw sequencing read counts and log2 transformed and DESeq normalized values formatted as genes-vs-samples matrix table .
|
| |
|
| Submission date |
Aug 21, 2020 |
| Last update date |
Aug 22, 2020 |
| Contact name |
Satish Kumar |
| E-mail(s) |
[email protected]
|
| Phone |
+1-956-665-6477
|
| Organization name |
University of Texas Rio Grande Valley
|
| Department |
Division of Human Genetics & South Texas Diabetes and Obesity Institute, UTRGV School of Medicine
|
| Street address |
5300 North L Street (MBMRF-2.219)
|
| City |
McAllen |
| State/province |
Texas |
| ZIP/Postal code |
78504 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (1) |
| GSE156617 |
Genome wide mRNA sequencing of induced pluripotent stem cell (iPSC) diffrentiated neural stem cells (NSCs). |
|
| Relations |
| BioSample |
SAMN15872635 |
| SRA |
SRX8983533 |
