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Status |
Public on Nov 26, 2008 |
Title |
NTHi 2h; 1740174013_B |
Sample type |
RNA |
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|
Source name |
Lung homogenate, 2h NTHi treatment
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Organism |
Mus musculus |
Characteristics |
C57BL/6
|
Treatment protocol |
The NTHi cell pellet was washed and resuspended in 20 ml 0.9% sodium chloride solution. This suspension was passed three times through an EmulsiFlex C5 cell disruptor (Avestin) at greater then 10,000 psi, then diluted to 4-5 mg/ml in 0.9% sodium chloride solution by bicinchoninic assay (Pierce) and centrifuged at 15,000 x g for 10 min. The supernatant was collected, the protein concentration was adjusted to 2.5 mg/ml, and the lysate was sterilized by passage through a 0.2 µm filter and frozen in 8 ml aliquots at -80ºC. For treatment, a thawed aliquot was placed in an AeroMist CA-209 nebulizer driven by 10 l/min 5% CO2 in air for 20 min, resulting in aerosolization of 4 ml of lysate, with the protein concentration in residual lysate
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Growth protocol |
Frozen stock (1 x 107 CFU/ml) of non-typeable Haemophilus influenzae (NTHi) was grown on chocolate agar at 300 ul per 10 cm plate (Remel) for 24 h at 37ºC in 5% CO2, then harvested and incubated for 16 h in 1 l brain-heart infusion broth (Acumedia) supplemented with 3.5 ug/ml NAD (Sigma-Adrich).
|
Extracted molecule |
total RNA |
Extraction protocol |
total RNA extracted using Qiagen Rneasy per manufacturers instructions. cRNA amplification using Ambion total prep kit per manufacturers instructions.
|
Label |
Cy3
|
Label protocol |
labeling with Cy3-Streptavidin per Illumina instructions.
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|
|
Hybridization protocol |
cRNA was hybridized to BeadChips per Illumina instructions.
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Scan protocol |
Chips were scanned using level 3 signal using a BeadStation 500 scanner.
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Description |
Gene expression in C57BL/6 mouse lungs after exposure to aerosolized NTHi lysate
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Data processing |
Data was processed using Illumina BeadStudio v3. Data generated both with rank invariant normalization, using sham group as the reference group.
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Submission date |
Nov 25, 2008 |
Last update date |
Nov 25, 2008 |
Contact name |
Scott E Evans |
E-mail(s) |
seevans@mdanderson.org
|
Organization name |
University of Texas-M.D. Anderson Cancer Center
|
Department |
Pulmonary Medicine - Unit 1462
|
Street address |
1400 Holcombe Blvd
|
City |
Houston |
State/province |
TX |
ZIP/Postal code |
77030 |
Country |
USA |
|
|
Platform ID |
GPL6105 |
Series (1) |
GSE13740 |
Gene expression in C57BL/6 mouse lungs after exposure to aerosolized NTHi lysate |
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