|
| Status |
Public on Apr 30, 2015 |
| Title |
RNA-Seq from Neural Stem Cells |
| Sample type |
SRA |
| |
|
| Source name |
primary neural stem cells
|
| Organism |
Mus musculus |
| Characteristics |
strain: BDF2
|
| Treatment protocol |
IES6 sample was generated by adding dox and 4OHT to the medium from day 1- 6
|
| Growth protocol |
Intermediate ES like cells (IES 6 cells): Nanog-CreER Reporter MEFs were plated at day 0 in MEF medium: DMEM supplemented with 1% L-Glutamine, 1% Non-essential amino acids, 1% penicillin/streptomycin and 10% FBS. On day 1 medium was changed to knock-out DMEM medium (Gibco 10829-018) with 10% KSR, 5% FBS, 1% L-Glutamine, 1% Non-essential amino acids, 1% penicillin/streptomycin, 0.05 mM β-mercaptoethanol and Dox+4OHT were added for a period of 6 days. By the end of day 6 tdTomato positive cells were sorted out using FACS sorter and used for RNA production. Primary Brain NSC: BDF2 mouse embryos were sacrificed at E13.5 of gestation according to standard protocols, and brain vesicles were mechanically dissociated and plated on Poly-D-Lysine and Laminin coated plates and were cultured in NSC medium: DMEM/F12 supplemented with 1XB27 and 1XN2 cell-supplements, 0.01% Fraction V BSA 7.5 % solution (Life Technologies), 1% Penicillin-streptomycin and 20ng/mL bFGF and 20ng/mL EGF.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were harvested and RNA was extracted using the 5 PRIME PerfectPure RNA Cell culture Kit (cat#2302340 ), according to manufacturer's manual
RNA was extracted from Trizol pellets, and utilized for RNA-Seq by TruSeq RNA Sample Preparation Kit v2 (Illumina) according to manufacturer’s instruction
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 1500 |
| |
|
| Description |
Brain derived NSC from E12.5 embryo
|
| Data processing |
We used Illumina CASAVA 1.8.2 software to generate fastq files.
We aligned reads to mm10 reference genome (index downloaded from bowtie) using TopHat version 2.0.10.
Read counts per exon were calculated over all 628,052 exons in mm10 ensemble GTF, using bedtools coverage command
Exon counts were normalized by the exon length in Kbp and by million number of aligned reads per sample, to give RPKM values.
Gene expression was defined by the average expression level (RPKM) of all exons associated with a certain gene.
Genome_build: mm10
Supplementary_files_format_and_content: exons.txt contains RPKM values calculated per-exon in all samples.
|
| |
|
| Submission date |
Mar 25, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Noa Novershtern |
| E-mail(s) |
noa.novershtern@weizmann.ac.il
|
| Organization name |
Weizmann Institute of Science
|
| Department |
Molecular Genetics
|
| Street address |
Weizmann Institute
|
| City |
Rehovot |
| ZIP/Postal code |
7610001 |
| Country |
Israel |
| |
|
| Platform ID |
GPL18480 |
| Series (2) |
| GSE67265 |
Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-Differentiation with iPSCs Reprogramming Factors (RNA-Seq) |
| GSE67299 |
Frequent and Transient Acquisition of Pluripotency During Somatic Cell Trans-Differentiation with iPSCs Reprogramming Factors |
|
| Relations |
| BioSample |
SAMN03447982 |
| SRA |
SRX969065 |
