|
| Status |
Public on Nov 02, 2022 |
| Title |
NA_KCl_0h_rep2_RNA_BS_seq |
| Sample type |
SRA |
| |
|
| Source name |
E16.5 mouse cortical neurons
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
kcl treatment: 0 h
developmental stage: E16.5
tissue/cell type: E16.5 mouse cortical neurons
genotype: WT
gender: pooled male and female
|
| Treatment protocol |
At DIV6, neuronal cells were silenced with 1 μM tetrodotoxin (TTX; Fisher) and 100 μM DL-2-amino-5-phosphopentanoic acid (DL-AP5; Fisher) overnight. The next morning, neuronal cells were depolarized with 55mM KCl for 0h, 2h, and 6h. At the end time point, the neuronal cells were harvested for RNA extraction.
|
| Growth protocol |
E16.5 mouse cortex tissues were dissected and dissociated into single cell suspension, resuspended in neuronal culture media (Neurobasal medium containing 2% B27 supplement (Invitrogen), 1% Glutamax (ThermoFisher) and 1% penicillin-streptomycin (ThermoFisher)) and seeded on laminin and poly-ornithine coated 10-cm dishes. Neurons were grown in vitro for 7 days with fresh medium changed on DIV3 and DIV6.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Total RNA was extracted from the neuronal cultures using TRIzol reagent combined with RNeasy min kit (QIAGEN) with DNase I on-column digestion.
After two rounds of poly(A) selection, bisulfite conversion was performed and RNA-seq libraries were prepared for sequencing using standard Illumina protocols
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Data processing |
Fastp v0.20 was used to remove adapter sequences, identify and remove low-quality reads and called bases
meRanGh v1.2.1 was used to map reads to C2T and G2A converted reference genomes
meRanCall was used to call m5C sites from mapped read files
In-house scripts using Python 3.9 and R 4.1.1 were used to further filter and process methylation calling files
Assembly: mm10
Supplementary files format and content: tab-delimited files containing methylation calling information for each called methylated site
Library strategy: RNA BS-Seq
|
| |
|
| Submission date |
Apr 26, 2022 |
| Last update date |
Nov 02, 2022 |
| Contact name |
Xiguang Xu |
| E-mail(s) |
xiguang@vt.edu
|
| Organization name |
Virginia Tech
|
| Department |
Department of Biomedical Sciences and Pathobiology
|
| Lab |
Epigenomics and Computational Biology Laboratory
|
| Street address |
1015 Life Science Circle
|
| City |
Blacksburg |
| State/province |
VA |
| ZIP/Postal code |
24060 |
| Country |
USA |
| |
|
| Platform ID |
GPL21103 |
| Series (2) |
| GSE201635 |
RNA bisulfite sequencing of mRNAs from E16.5 mouse cortical neurons depolarized with 55mM KCl for 0h, 2h and 6h. |
| GSE201637 |
RNA bisulfite sequencing and RNA-seq of 16.5 mouse cortical neurons depolarized with 55mM KCl for 0h, 2h and 6h |
|
| Relations |
| BioSample |
SAMN27861737 |
| SRA |
SRX15010766 |
