|
| Status |
Public on Nov 01, 2015 |
| Title |
A549_TULV_12h_1 |
| Sample type |
RNA |
| |
|
| Source name |
A549_TULV_12h
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: A549
inoculum: TULV Moravia Ve6P3951129
|
| Treatment protocol |
Cells were infected with the described viruses at MOI of 5.0 and the virus was adsorbed for 1h. Mock infections were performed using culture medium free of any virus.
|
| Growth protocol |
A549 human lung carcinoma epithelial cells (DSMZ ACC 107) were cultured under standard conditions in 12-well cell culture plates
|
| Extracted molecule |
total RNA |
| Extraction protocol |
TRIzol® extraction, according to manufacturer's protocol
|
| Label |
Cy3
|
| Label protocol |
For the linear T7-based amplification step, 100 ng of each total RNA sample was used. To produce Cy3-labeled cRNA, the RNA samples were amplified and labeled using the Agilent Low Input Quick Amp Labeling Kit (Agilent Technologies) following the manufacturer’s protocol. Yields of cRNA and the dye-incorporation rate were measured with the ND-1000 Spectrophotometer (NanoDrop Technologies).
|
| |
|
| Hybridization protocol |
The hybridization procedure was performed according to the Agilent 60-mer oligo microarray processing protocol using the Agilent Gene Expression Hybridization Kit (Agilent Technologies). Briefly, 600 ng Cy3-labeled fragmented cRNA in hybridization buffer was hybridized overnight (17 hours, 65 °C) to Agilent Whole Human Genome Oligo Microarrays 8x60K using Agilent’s recommended hybridization chamber and oven. Finally, the microarrays were washed once with the Agilent Gene Expression Wash Buffer 1 for 1 min at room temperature followed by a second wash with preheated Agilent Gene Expression Wash Buffer 2 (37 °C) for 1 min. The last washing step was performed with acetonitrile.
|
| Scan protocol |
Fluorescence signals of the hybridized Agilent Microarrays were detected using Agilent’s Microarray Scanner System (Agilent Technologies).
|
| Data processing |
The Agilent Feature Extraction Software (FES) was used to read out and process the microarray image files. For determination of differential gene expression FES derived output data files were further analyzed using the Rosetta Resolver gene expression data analysis system (Rosetta Biosoftware).
|
| |
|
| Submission date |
Sep 24, 2015 |
| Last update date |
Nov 01, 2015 |
| Contact name |
Daniel Bourquain |
| E-mail(s) |
bourquaind@rki.de
|
| Phone |
0049 (0)30187543825
|
| Organization name |
Robert Koch Institute
|
| Department |
Centre for Biological Threats and Special Pathogens 1
|
| Lab |
Highly Pathogenic Viruses
|
| Street address |
Seestraße 10
|
| City |
Berlin |
| State/province |
Berlin |
| ZIP/Postal code |
13353 |
| Country |
Germany |
| |
|
| Platform ID |
GPL13607 |
| Series (1) |
| GSE73410 |
Diversity in Modulation of Interferon Stimulated Genes by Dobrava, Kurkino, and Sochi Genotypes of Dobrava-Belgrade Hantavirus in comparison to Tula Virus |
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