|
| Status |
Public on Oct 17, 2013 |
| Title |
SVG 5F4 Clone Mock Infected Replicate 3 |
| Sample type |
RNA |
| |
|
| Source name |
SVG Clone 5F4
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: SVG
clone: 5F4
infection: Mock
jcv replication rate: NA
time post infection: 14 days
|
| Treatment protocol |
Cells were allowed to attach to the wells of 6 well dishes at a density of 1.5x105 cells per well. Approximately 24 hours after seeding, cells were exposed to 400 HAU of Mad-4 JCV per 1x106 cells or medium alone for the mock condition. Acutely infected cells were fed with new medium at 7 days post-infection. Samples were collected at 0, 7, and 14 days to monitor infection. RNA from 14 days post-infection or mock infection was used for microarray analysis.
|
| Growth protocol |
SVG cells were previously generated by transfecting human fetal brain cultures with an origin-defective SV40 mutant, which was identical to the expression vector used to produce the COS cell lines. The resultant culture of multiple phenotypes of neural derived cells became immortalized by stable expression of SV40 T antigen. SVG cells can be maintained in media containing 10% serum, but to remove serum concentration as an experimental variable, SVG cells were maintained in the same media as SVG-derived clones: Eagle’s minimal essential medium EMEM supplemented with 20% FBS, 2 mM L-glutamine, and penicillin/streptomycin. Cells were grown at 37°C and 5% CO2. Cells were subcultured 1:4 when confluent.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted by sample using the Qiagen RNAeasy kit. Post extreaction, RNA quality and quantity was ensured using the Bioanalyzer (Agilent, Inc) and NanoDrop (Thermo Scientific, Inc) respectively.
|
| Label |
Biotin
|
| Label protocol |
200 nanograms of total RNA per sample was used in conjunction with the Affymetrix recommended protocol for the GeneChip Human Gene 1.0 ST Array (Affymetrix, Inc).
|
| |
|
| Hybridization protocol |
Labeled cDNA was hybridized, washed and stained to separate Affymetrix GeneChip Human Gene 1.0 ST Arrays by sample according to the protocols described by Affymetrix (Affymetrix, Inc). Per staining, streptavidin phycoerythrin solution was used (Molecular Probes, Carlsbad, CA) and enhanced by an antibody solution containing 0.5 mg/mL of biotinylated anti-streptavidin (Vector Laboratories, Burlingame, CA).
|
| Scan protocol |
The Affymetrix Gene Chip Scanner 3000 was used to scan each Affymetrix GeneChip Human Gene 1.0 ST Array. After, gene probe intensities were generated using the Affymetrix AGCC software (Affymetrix, Inc).
|
| Description |
SVG 5F4 Clone Mock Infected Replicate 3
|
| Data processing |
The Affymetrix Expression Console (Affymetrix, Inc) was used to summarize the gene probe intensities and generate 33,297 RMA normalized gene fragment expression values for each hybridized cRNA.
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| |
|
| Submission date |
Mar 29, 2013 |
| Last update date |
Oct 18, 2013 |
| Contact name |
Kory R Johnson |
| E-mail(s) |
[email protected]
|
| Phone |
301-402-1956
|
| Organization name |
NINDS/NIH
|
| Department |
DIR IT & Bioinformatics
|
| Lab |
Bioinformatics Section
|
| Street address |
10/3B01, 9000 Rockville Pike
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL6244 |
| Series (1) |
| GSE45639 |
Clonal Immortalized Human Glial Cell Lines Support Varying Levels of JC Virus Infection due to Differences in Cellular Gene Expression |
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