|
| Status |
Public on Sep 05, 2015 |
| Title |
microRNA IM103 VN1203 4d RNA 5 |
| Sample type |
RNA |
| |
|
| Source name |
Lung tissue, VN1203-inoculated, 4 day(s), bioreplicate 5
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57Bl/6J
age: 22 weeks
time: 4
virus: VN1203
biological_replicate: 5
|
| Treatment protocol |
Mice were anesthetized with isofllurane and inoculated with PBS or PBS containing virus in a volume of 50 μl.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from TRIzol homogenate of mouse lung.
|
| Label |
Cy3
|
| Label protocol |
One-color labeling reactions were prepared using the Agilent One-Color Protocol (Version 6.6). 200 ng of total RNA with the appropriate concentration of RNA spike-in controls is converted into double stranded cDNA using an olido (dT) primer linked to the T7 promoter sequence and MMLV-RT. The double stranded cDNA is then in vitro transcribed by T7 RNA polymerase, which simultaneously amplifies and incorporates cyanine 3-labeled CTP into the resulting cRNA. Labeled cRNA is then purified using Qiagen RNeasy columns and the labeling efficiency determined by the NanoDrop spectrophotometer.
|
| |
|
| Hybridization protocol |
The labeled cRNA is fragmented and placed on the mouse 8x60K Gene Expression microarray and hybridized at 65°C for ~17 hours.
|
| Scan protocol |
The hybridized arrays are washed and scanned at 3 μM resolution on an Agilent G2565CA High Resolution Scanner.
|
| Data processing |
Scanned images were analyzed using Agilent Feature Extraction Software (v11.5.1.1) using the protocol GE1_1105_Oct12. The limma package for R (available on Bioconductor) was used to perform background correction, quantile normalization (normalizeBetweenArrays), and summarization (avereps) to derive a single normalized intensity value per probe. Outlier samples were detected using PCA and by visual inspection of heatmaps, and all data was re-processed after removing outlier samples. All data processing for each of the biological replicates was performed independently of the other.
|
| |
|
| Submission date |
Aug 25, 2015 |
| Last update date |
Aug 31, 2016 |
| Contact name |
Natalie Heller |
| E-mail(s) |
natalie.heller@pnnl.gov
|
| Organization name |
PNNL
|
| Street address |
902 Battelle Blvd.
|
| City |
Richland |
| ZIP/Postal code |
99354 |
| Country |
USA |
| |
|
| Platform ID |
GPL19970 |
| Series (2) |
| GSE65575 |
Modeling Host Responses to Understand Severe Human Virus Infections |
| GSE72365 |
Mouse lung tissue transcriptome response to influenza H5N1 [miRNA] (AmbryGenetics) |
|
