|
| Status |
Public on Jan 20, 2016 |
| Title |
AdGFP_48hr_3 |
| Sample type |
SRA |
| |
|
| Source name |
Cultured Primary Hepatocytes
|
| Organism |
Rattus norvegicus |
| Characteristics |
cell type: hepatocyte
strain: Sprague-Dawley
treatment: AdGFP-infected
collection time: 48h post-plating/24h post-infect
|
| Treatment protocol |
Cells were left uninfected over 72hr or 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 the sequencing facility of the Drexel University College of Medicine Center for Genomic Sciences using the Illumina TruSeq Stranded mRNA prep kit
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Sequencing and base calling were done using an Illumina NextSeq following standard methods by 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
Supplementary_files_format_and_content: Tab-delimited with header. Each row has counts/rpkm for all samples of each gene feature in the Rn5 GTF annotation (named in first row).
|
| |
|
| Submission date |
Apr 21, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Michael Bouchard |
| E-mail(s) |
michael.bouchard@drexelmed.edu
|
| Organization name |
Drexel University College of Medicine
|
| Department |
Biochemistry
|
| Street address |
245 N 15th St
|
| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19102 |
| Country |
USA |
| |
|
| Platform ID |
GPL20084 |
| Series (2) |
| GSE68112 |
Hepatitis B virus-mediated alterations to the primary hepatocyte transcriptome [Full Set] |
| GSE68113 |
Hepatitis B virus-mediated alterations to the primary hepatocyte transcriptome |
|
| Relations |
| SRA |
SRX1002908 |
| BioSample |
SAMN03497697 |
