|
| Status |
Public on Apr 30, 2019 |
| Title |
1hr_NoAb_replicate2_QM5 |
| Sample type |
RNA |
| |
|
| Source name |
BMDM, NoAb, 1hr
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Macrophages, bone marrow-derived, from female C57B/6 mice from Jackson Labs
slide id (array batch): 257480911094
time: 1hr
treatment: NoAb
|
| Treatment protocol |
BMDMs were seeded in 6-well dishes at 1xE06 cells per well. Cells were activated overnight with IFNγ and LPS. Cells were then treated for 1 h with or without Fc receptor blocking antibody, or harvested before the incubation for a baseline. Cells were lifted from the dishes, pelleted, and resuspended in TRIzol reagent [ThermoFisher]. The supernatant was then flash frozen and stored at −80 °C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the PureLink RNA Mini kit (Ambion/Life Technologies) according to the manufacturer’s protocol, including on-column DNAse treatment. Following elution of purified RNA from the PureLink columns with Nuclease-free water, quantitation was performed using a NanoDrop spectrophotometer and quality assessment determined by RNA LabChip analysis on an Agilent BioAnalyzer 2100 or RNA Screen tape on an Agilent TapeStation 2200.
|
| Label |
Cy3
|
| Label protocol |
One hundred nanograms of total RNA was processed for hybridization to Agilent SurePrint G3 Mouse v2 8x60K Gene Expression Arrays according Agilent’s One Color Microarray-Based Analysis (Low Input QuickAmp Labeling) protocol, including cDNA synthesis, cRNA synthesis with Cy3 labeling and purification, fragmentation, hybridization, and post-hyb washing. Spike-In controls were utilized and processed according to Agilent’s One Color RNA Spike-In kit protocol.
|
| |
|
| Hybridization protocol |
600ng of Cy3-labelled cRNA was purified, fragmented and hybridized to Agilent SurePrint G3 Mouse v2 8x60K Gene Expression Arrays, according to the Agilent Low Input QuickAmp Labeling Kit protocol. Post-hybridization washing of arrays was also performed according to these protocols.
|
| Scan protocol |
The arrays were scanned in an Agilent G2600D SureScan Microarray Scanner using scan protocol AgilentG3_GX_1color. Agilent’s Feature Extraction Software was used to assign grids, provide raw image files per array, and generate QC metric reports from the microarray scan data. The QC metric reports were used for quality assessment of all hybridizations and scans.
|
| Description |
Gene Expression Data
|
| Data processing |
Txt files from the Feature Extraction software were imported into the Partek Genomics Suite (v7.0; Partek) for detailed analyses of gene expression. Within Partek, the gProcessedSignal was imported and the intensity values were normalized to the 75th percentile, lower expressed genes were filtered out, and log transformation base 2.0 was performed. The batch effect removal tool (analysis of variance [ANOVA]) was used to correct for effects of array (slide).
|
| |
|
| Submission date |
Feb 23, 2019 |
| Last update date |
May 01, 2019 |
| Contact name |
Anne E. Jedlicka |
| Organization name |
Johns Hopkins University, School of Public Health
|
| Department |
Molecular Microbiology and Immunology
|
| Lab |
Genomic Analysis and Sequencing Core
|
| Street address |
615 N. Wolfe street
|
| City |
Baltimore |
| State/province |
MD |
| ZIP/Postal code |
21205 |
| Country |
USA |
| |
|
| Platform ID |
GPL21810 |
| Series (1) |
| GSE126977 |
Dragotcytosis: Elucidation of the Mechanism for Cryptococcus neoformans Macrophage-to-Macrophage Transfer |
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