|
| Status |
Public on Mar 08, 2021 |
| Title |
Control IgG 1 |
| Sample type |
RNA |
| |
|
| Source name |
whole lung homogenate
|
| Organism |
Mus musculus |
| Characteristics |
tissue: whole lung
strain: C57/BL6
treatment: control IgG and E. coli
age: 10-12 weeks
|
| Treatment protocol |
After anesthetization with ketamine and xylazine, the left lung lobes of C57Bl/6J mice were intratracheally infected with 1E6 CFU E. coli plus either 10 ng IgG control antibody or neutralizing anti-OSM antibody. After 6 hours of infection, whole left lung lobes were harvested, frozen at -80˚C and stored for future RNA isolation.
|
| Growth protocol |
Male and female C57BL/6J mice were obtained from Jackson Laboratories at 6-7 weeks of age. Prior to infection, they were housed at the Boston University animal facility and provided with an ad libitum supply of standard chow. Mice were infected at 10-12 weeks of age. All animal protocols were approved by the BU Institutional Animal Care and Use Committee (IACUC).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Frozen left lobes in RLT buffer were homogenized using a Bullet Blender (Next Advance, Troy, NY) per the manufacturer’s instruction. Cellular debris was removed by centrifugation (max speed 3 minutes). RNA was isolated using the RNeasy micro kit (Qiagen), following the manufacturer’s instructions. RNA was eluted in 50µl RNase-free water.
|
| Label |
biotin
|
| Label protocol |
Biotin labeling was performed using the WT Plus reagent kit (Affymetrix, Santa Clara, CA) according to the manufacturer's protocol.
|
| |
|
| Hybridization protocol |
The labeled, fragmented DNA was hybridized to the Affymetrix Mouse Gene 1.0 ST Array for 18 hours in a GeneChip Hybridization oven 640 at 45oC with rotation (60 rpm). The hybridized samples were washed and stained using an Affymetrix fluidics station 450.
|
| Scan protocol |
After staining, microarrays were immediately scanned using an Affymetrix GeneArray Scanner 3000 7G Plus.
|
| Description |
Gene expression data from whole lung homogenate of C57/BL6 mice treated with control IgG and E. coli.
|
| Data processing |
Raw Affymetrix CEL files were normalized to produce Entrez Gene-identifier-specific expression values using the implementation of the Robust Multiarray Average (RMA) in the affy Bioconductor package (version 1.36.1), using R version 2.15.1 and the Brainarray mogene10stmmentrezgcdf R package (version 16.0.0).
|
| |
|
| Submission date |
Jul 28, 2020 |
| Last update date |
Mar 09, 2021 |
| Contact name |
Boston University Microarray and Sequencing Resource |
| E-mail(s) |
[email protected]
|
| Organization name |
Boston University
|
| Department |
Microarray and Sequencing Resource
|
| Street address |
72 East Concord Street, E631
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02118 |
| Country |
USA |
| |
|
| Platform ID |
GPL19485 |
| Series (1) |
| GSE155283 |
Oncostatin M neutralization in E. coli pneumonia |
|
