|
| Status |
Public on Jan 15, 2016 |
| Title |
gfp_48_2 |
| Sample type |
SRA |
| |
|
| Source name |
Cultured Primary Hepatocytes
|
| Organism |
Rattus norvegicus |
| Characteristics |
cell type: hepatocytes
strain: Sprague-Dawley
treatment: AdGFP-infected
collection time: 48h post-infection/72h post-plating
|
| Treatment protocol |
Cells were infected with either AdGFP or AdHBV 24h after isolation and plating. 24h or 48h post-infection cells were washed with 1X PBS and stored at -80° until RNA isolation
|
| Growth protocol |
Primary rat hepatocytes were isolated using a 2-step perfusion protocol (Seglen, P. 1993. Methods Toxicol.) and plated on collagen coated 6cm plates. Cells were maintained in Williams E Medium supplemented with 2 mM L-glutamine, 1 mM sodium pyruvate, 4 μg/ml insulin-transferrin-selenium, 5 μg/ml hydrocortisone, and 5 ng/ml epidermal growth factor at 37°C in 5% CO2. Medium was changed on cells daily.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Total RNA was isolated using the mirVana RNA isolation kit following the manufacturer's total RNA isolation protocol. Prior to cDNA library prep RNA was polyA selected
Libraries were prepared by GENEWIZ, Inc. using the NEBNext Ultra RNA Library Prep following the manufacturer's protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
Sequencing and base calling were done using an Illumina HiSeq following standard methods by GENEWIZ, Inc. (1x50bp reads) or with an Illumina NextSeq 500 instrument at the Drexel University College of Medicine Center for Genomic Sciences (1x75bp reads)
Reads were aligned to a reference genome consisting of the Rn5 build of the rat genome, the HBV serotype ayw sequence, the hrGFP sequence, and the pAdEasy1 sequence using the STAR aligner (v2.4.0h) using the options --outSAMtype BAM SortedByCoordinate and --outFilterMismatchNoverLmax 0.05
Mapped reads were filtered for "mapping quality = 255" using Sambamba (v0.4.7)
Reads per kilobase feature per megabase of library (RPKM) was calculated by counting reads per feature using the GenomicAlignments package (v1.2.1) in R (v3.1.2) and dividing these counts by kilobase of feature length to get RPK, then dividing by millions of reads per sample to get RPKM.
genome build: Rnor_5.0
processed data files format and content: Tab-delimited with header. Each row has counts/rpkm for all 12 samples from merged BAM files) of each gene feature in the Rn5 GTF annotation (named in first row).
|
| |
|
| Submission date |
Mar 05, 2015 |
| Last update date |
Oct 07, 2024 |
| Contact name |
James M Wilson |
| Organization name |
University of Pennsylvania
|
| Street address |
125S 31st St
|
| City |
Philadelphia |
| State/province |
Pennsylvania |
| ZIP/Postal code |
19104 |
| Country |
USA |
| |
|
| Platform ID |
GPL18694 |
| Series (2) |
| GSE66566 |
Hepatitis B virus-mediated alterations to the primary hepatocyte transcriptome [24 and 48 h] |
| GSE68113 |
Hepatitis B virus-mediated alterations to the primary hepatocyte transcriptome |
|
| Relations |
| BioSample |
SAMN03389395 |
| SRA |
SRX901612 |
