|
| Status |
Public on Nov 02, 2014 |
| Title |
RNA-seq from Mettl3-KO ESCs, replicate B |
| Sample type |
SRA |
| |
|
| Source name |
Embryonic Stem Cells
|
| Organism |
Mus musculus |
| Characteristics |
genotype: Mettl3-KO
cell type: Embryonic Stem Cells
strain: C57BL/6
|
| Growth protocol |
Maintenance of grounds state naïve murine pluripotent cells was conducted in FBS free N2B27-based media, and included 10μg recombinant human LIF and small-molecule inhibitors CHIR99021 (CH, 1-3 μM- Axon Medchem) and PD0325901 (PD, 1 μM - TOCRIS) termed 2i. For embryoied bodies formation 5x10^6 ESCs were disaggregated with trypsin and transfer to low non-adherent palte and culture in MEF medium (DMEM supplemented with 1% L-Glutamine, 1% Non-essential amino acids, 1% penicillin/streptomycin and 15% FBS) for 8-21 days. MEF generation: E13.5 mouse embryo was decapitated and disaggregated with trypsin and culture in MEFs medium until MEFs population were expended.
|
| Extracted molecule |
polyA 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
Libraries were prepared according to Illumina's instructions accompanying the TruSeq RNA Sample Preparation Kit v2 ( RS-122-2001).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| 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.
To estimate expression levels of all genes we used HTSeq-count software, version 0.6.0 ,that was ran with mm10 gtf file downloaded from ensemble.
To get normalized expression levels (FPM) we used DESeq R library
Genome_build: mm10
|
| |
|
| Submission date |
Oct 02, 2014 |
| 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 |
GPL17021 |
| Series (2) |
| GSE61997 |
m6A mRNA Methylation Facilitates Resolution of Naïve Pluripotency Towards Differentiation (RNA-Seq) |
| GSE61998 |
m6A mRNA Methylation Facilitates Resolution of Naïve Pluripotency Towards Differentiation |
|
| Relations |
| Reanalyzed by |
GSE80797 |
| BioSample |
SAMN03092742 |
| SRA |
SRX719379 |
