|
| Status |
Public on Nov 21, 2012 |
| Title |
gmcsf ko water 6 days 78_2 |
| Sample type |
RNA |
| |
|
| Source name |
colon crypts from untreated gmcsf ko mice
|
| Organism |
Mus musculus |
| Characteristics |
genotype: GMCSF KO
treatment: water
gender: male
tissue: colon crypts
strain: C57BL/6
age: 8 weeks
|
| Treatment protocol |
Colitis was induced by allowing mice to drink 1.5% (w/v) Dextran sodium sulfate (DSS) (m. w. 36-50 kDa, MP Biomedicals) dissolved in the drinking water ad libitum for 6 days. Untreated mice were administered water only.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA (isolated by RNeasy kit; Qiagen following the manufacturer's intructions) was prepared from isolated crypts of WT and GM-CSF-/- mice colons before or after 1.5% DSS treatment for 6 days. .
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 150ng RNA using the Quick Amplification Kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
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| |
|
| Hybridization protocol |
1650ng of Cy3-labelled cRNA was fragmented at 60°C for 30 minutes in a reaction volume of 55ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 55ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Mouse Genome Microarray Kit, 4x44K (G4122F) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B). 5 µm scanning in green at 10% and 100% PMT
|
| Data processing |
Slide design ID was 26655 and the protocol used for extraction was GE1_107_Sep09. Version 10.7.3.1. of feature extraction. The data were normalized using the multi-loess method described in this paper: Microarray truths and consequences, R. Sasik, C. H. Woelk and J. Corbeil, J. Mol. Endocrinology, 33, 1 (2004)
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| |
|
| Submission date |
Nov 08, 2012 |
| Last update date |
Nov 21, 2012 |
| Contact name |
Laia Egea |
| E-mail(s) |
legeapujol@ucsd.edu
|
| Organization name |
UCSD
|
| Department |
Medicine
|
| Lab |
Laboratory of Mucosal Immunology
|
| Street address |
9500 Gilman Dr
|
| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92093-0623 |
| Country |
USA |
| |
|
| Platform ID |
GPL10333 |
| Series (1) |
| GSE42173 |
Expression profile of colon crypts from WT and GMCSF-/- mice with or without DSS treatment |
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