|
| Status |
Public on Sep 01, 2021 |
| Title |
ST_Valproic_10000_b1_p4_D06 |
| Sample type |
SRA |
| |
|
| Source name |
liver
|
| Organism |
Rattus norvegicus |
| Characteristics |
strain: Wistar
gender: Male
cell type: hepatocytes
tempo-seq assay: S1500+ rat surrogate assay
chemical: Valproic
dose (um): 10000
batch_id: 1
plate_id: 4
well_id: D06
|
| Treatment protocol |
Chemical treatments were performed in culture medium containing 0.5 % DMSO for 24 h.
|
| Growth protocol |
Freshly isolated hepatocytes were maintained in collagen sandwich at 25,000 cells/well in a 96-well plate with William’s medium E containing 1 mg/mL BSA, 0.5 µg/mL insulin, 2 mM L-glutamine, 100 nM dexamethasone, 10 ng/mL EGF, 100 μg/mL streptomycin and 100 U/mL penicillin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
After 24 h of chemical treatment, the spent medium was removed and the wells were washed with 100 µL of PBS twice. Cells were lysed by adding 20 µL of 1X TempO-Seq lysis buffer (BioSpyder) in PBS. The cell lysates were shipped frozen on dry ice to to BioSpyder Technologies, Inc. (Carlsbad, CA, USA), where libraries were prepared directly from the cell lysates and RNA-seq was performed.
2 μL of sample lysate was hybridized with detector oligos from the TempO-Seq rat surrogate (ST) or whole-transcriptome (WT) assay, with 1 h hybridization period for the ST assay and overnight for the WT assay. After ligation and nuclease digestion, the ligated products were added into an amplification mix with sample-specific PCR primer pairs which include the standard Illumina adaptor and a sample-specific barcode sequence. Amplicons were pooled and purified using a PCR clean-up kit (Macherey-Nagel, Mountain View, CA, United States).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
ST pilot combined FINAL.csv
|
| Data processing |
Data processing was performed by BioSpyder Technologies Inc.
Illumina Casava1.7 software used for base-calling
No filtering was done since aligning to known sequence
Reads were aligned using the STAR aligner 2.5.3a allowing 2 bp mismatches and not allowing indels. The softclipping option is turned on.
The Tempo-SeqR package was used to generate read counts matrix
Genome_build: rn6
Supplementary_files_format_and_content: Read counts matrix (row = treatment; column = gene)
|
| |
|
| Submission date |
Jun 09, 2020 |
| Last update date |
Sep 01, 2021 |
| Contact name |
Fan Lee |
| Organization name |
Institute of Bioengineerng and Nanotechnology
|
| Lab |
Prof Hanry Yu
|
| Street address |
31 Biopolis Way, The NANOS, #06-01
|
| City |
Singapore |
| ZIP/Postal code |
138669 |
| Country |
Singapore |
| |
|
| Platform ID |
GPL18694 |
| Series (1) |
| GSE152128 |
High-throughput transcriptomics platform for screening hepatotoxicants |
|
| Relations |
| BioSample |
SAMN15189467 |
| SRA |
SRX8510817 |
