|
| Status |
Public on Jul 29, 2019 |
| Title |
Neurons5_Prep1 |
| Sample type |
SRA |
| |
|
| Source name |
single-cell CVB derived hiPSC#5 subline (genome editing: PSEN1 deltaE9 -/-, IIA12)
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Craig Venter-B (CVB) hiPSC subline
cell type: single-cell derived from CVB line prior to neuronal differentiation, preparation 1
|
| Treatment protocol |
No particular treatment was applied after differentiation.
|
| Growth protocol |
hiPSCs were seeded onto PA6 cells in PA differentiation medium for 12 days. Between days 12-14, when rosettes were visible, after dissociation, were FACS-sorted based on the CD184+/CD271-/CD44-/CD24+ cell-surface signature. Sorted NPCs were cultured on 20µg/mL poly-L-ornithine and 5µg/mL laminin coated plates in NPC medium until confluency was reached. To generate neurons, NPCs grown in 10 cm-plates at confluency were switched to NPC medium without FGF (day 1), and cultured afterwards for 3 weeks. Around day 21, neurons were sorted based on the CD184-/CD44-/CD24+ signature. For RNAseq experiments 1x106 CD24+ sorted neurons were plated on 20µg/mL poly-L-ornithine and 5µg/mL laminin coated 24-well plates in NPC media supplemented with 0.5mM dbcAMP from Sigma, and 20ng/µL BDNF and 20 ng/µL GDNF.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Total RNA was extracted using RNeasy Assay (Qiagen) and converted into cDNA using SuperScript First-Strand Synthesis System (Invitrogen) following manufacture’s protocols.
RNA quality was calculated on TapeStation (Agilent Technologies). Most of samples showed a RIN >7.0 and 7 out of 24 had an RIN between 6 and 7. PolyA+ RNA-seq libraries were generated using the Illumina TruSeq Stranded RNA LT kit and following manufacturer’s detailed instructions.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
Induced neuronal differentiation from hiPSCs
|
| Data processing |
Reads were aligned to the hg18 human genome with bowtie2 (–very-sensitive option) and tophat. One mismatch was allowed.
Artifacts derived from clonal amplification were circumvented by considering maximal three tags from each unique genomic position as determined from the mapping data
Read counts were calculated with HOMER 3.9 considering only exonic regions of human RefSeq genes
The bedGraph and bigwig files were generated by using Homer v3.9, which the total tags are normalized to 1.00e+07.
Genome_build: hg18
Supplementary_files_format_and_content: bedGraph
|
| |
|
| Submission date |
Nov 14, 2017 |
| Last update date |
Jul 30, 2019 |
| Contact name |
Daria Merkurjev |
| E-mail(s) |
[email protected]
|
| Phone |
858-534-5858
|
| Organization name |
UCSD
|
| Department |
Medicine
|
| Lab |
Michael G. Rosenfeld Laboratory
|
| Street address |
9500 Gilman Drive, Mail Code 0648
|
| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92093 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (2) |
| GSE106871 |
Analysis of the Clustered Protocadherin (cPcdh) Locus in Human Pluripotent Stem and Derived Cells (RNA-seq I out of II) |
| GSE106872 |
Analysis of the Clustered Protocadherin (cPcdh) Locus in Human Pluripotent Stem and Derived Cells |
|
| Relations |
| BioSample |
SAMN08024734 |
| SRA |
SRX3390517 |
