|
| Status |
Public on Aug 03, 2023 |
| Title |
D6_1_RNA |
| Sample type |
SRA |
| |
|
| Source name |
P19
|
| Organism |
Mus musculus |
| Characteristics |
cell line: P19
cell type: Embryonal carcinoma
genotype: WT
treatment: Neuronal differentiation
|
| Treatment protocol |
To induce neuronal differentiation, 3x105 cells were seeded in poly-L-lysine-coated 6-well plates in the presence TaKaRa NDiff® 227 Medium and 500 nM retinoic acid. Media was changed daily up to 6 days after induction.
|
| Growth protocol |
Mouse embryonal carcinoma P19 cells (ATCC) were maintained in Dulbecco's modified Eagle's medium with 4,500 mg/L of glucose supplemented with 100 units/mL penicillin/streptomycin (Life Technologies) and 10% fetal bovine serum (FBS, Gemini Bio-Products).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cell lysates from each sample were split to generate the RPF and RNA-seq libraries. Total RNA from the cell lysates used for the RNA-seq libraries preparation was extracted using Qiazol.
To generate the RPF libraries, the lysates were digested with Mnase. The lysates were run on a polyacrylamide-Urea gel, and fragments 20-40 nucleotides long were purified. The NEBnext library preparation kit for small RNAs was used to generate libraries from the purified RPFs fragments. Illumina RNA-seq libraries were generated from the total RNA extracted from the cell lysates.
Libraries were sequenced on a High Output Flow Cell by a NextSeq 500 sequencer (Illumina), utilizing a 51 bp single-end protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Base calling was done with the Illumina software bcl2fastq v2.20
Optical duplicates were removed using clumpify from the bbtools suite
Adapters were trimmed using bbduk from the bbtools suite
rRNA was removed using bbduk
Reads were filtered to within 20-50nt length (for RNA-Seq) or 26-34nt length (for RFP) using bbduk.
Reads were mapped using bbmap from the bbtools suite in ‘slow’ mode.
The differential translational engagement was determined with Xtail using read counts of RNA-seq and RFP libraries from D0 and D6 samples as input.
Assembly: mm10 assembly annotated with GENCODE vM23 genes
Supplementary files format and content: Table with differential translational efficiency between D0 and D6.
Library strategy: Ribo-seq
|
| |
|
| Submission date |
Oct 04, 2022 |
| Last update date |
Aug 03, 2023 |
| Contact name |
Marcos Morgan |
| Organization name |
NIEHS
|
| Street address |
111 TW Alexander Dr
|
| City |
Durham |
| ZIP/Postal code |
27709 |
| Country |
USA |
| |
|
| Platform ID |
GPL19057 |
| Series (2) |
| GSE214816 |
Differential ribosome engagement profiling between undifferentiated and neuron-differentiated P19 cells |
| GSE214875 |
Post-transcriptional regulation of Ribosome Biogenesis during differentiation |
|
| Relations |
| BioSample |
SAMN31155138 |
| SRA |
SRX17797436 |
