|
| Status |
Public on Dec 21, 2017 |
| Title |
m6A-seq Gsk-3 DKO |
| Sample type |
SRA |
| |
|
| Source name |
Embryonic stem cells
|
| Organism |
Mus musculus |
| Characteristics |
strain: E14K
antibody: m6A antibody, NEB, E1611A, lot 0011602
genotype/variation: Gsk-3 DKO
|
| Treatment protocol |
Cells were grown in the absence of LIF for 14 days before isolating total RNA using Trizol.
|
| Growth protocol |
Feeder-free ESCs were grown on 10 cm plates coated with 0.1 % gelatin. Media was high glucose DMEM supplemented with 15% FBS, 1% non-essential amino acids, 1% sodium pyruvate, 1% L-glutamine, 1% penicillin/streptomycin, and 55 μM β-mercaptoethanol.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Purified total RNA was then incubated with oligo (dT) beads to isolate mRNA. m6A mRNA was subsequently isolated using Dynabeads coupled with m6A antibody.
m6A mRNA and total mRNA was chemically fragmented by incubating with 100 mM Tris HCl, pH 7.0 and 100 mM ZnCl2 at 94°C for 5 minutes. 100 ng of RNA were used to prepare libraries according to Ion XpressRNA-seq barcoding kit protocols.
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Ion Torrent Proton |
| |
|
| Data processing |
Library strategy: m6A-seq
m6A-seq reads were aligned to the reference genome using Bowtie 2.
The DESeq package in StrandNGS software was used to quantify mRNA and identify mRNAs with differential amounts of m6A between WT and Gsk-3 DKO ESCs.
Genome_build: mm9
Supplementary_files_format_and_content: Peaks_with_gene_annotations_filtered_ERD_sorted
|
| |
|
| Submission date |
Dec 20, 2017 |
| Last update date |
Dec 21, 2017 |
| Contact name |
Christopher Phiel |
| E-mail(s) |
christopher.phiel@ucdenver.edu
|
| Phone |
303-315-7678
|
| Organization name |
University of Colorado Denver
|
| Department |
Integrative Biology
|
| Street address |
1150 12th Street SI 4111
|
| City |
Denver |
| State/province |
CO |
| ZIP/Postal code |
80204 |
| Country |
USA |
| |
|
| Platform ID |
GPL18635 |
| Series (1) |
| GSE108327 |
m6A-seq in wild-type and Gsk-3-deficient mouse embryonic stem cells |
|
| Relations |
| BioSample |
SAMN08213141 |
| SRA |
SRX3489014 |
