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GEO help: Mouse over screen elements for information. |
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Status |
Public on Dec 21, 2005 |
Title |
Expression microarray analysis of acquired gemcitabine resistance |
Organism |
Mus musculus |
Experiment type |
Expression profiling by array
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Summary |
This series repressents the data set from the paper "Expression microarray analysis and oligo array CGH of aquired gemcitabine resistance in mouse colon reveals selection for chromosomal aberrations".
Biological material: One solid mouse tumor "Colon 26A" was routinely maintained by successive transplantation. A subset of mice with this tumor was treated at the maximum tolerable dosage of Gemcitabine and successively transplanted 5 times and similarly treated. The last passage was completly resistant and designated, "Colon 26G".
Expression array: Total RNA was isolated from frozen mouse tumors using Tryzol. Single stranded cDNA was synthetized from 30 mg of total RNA by reverse transcription using aminoallyl labeled dUTP (Ambion). Labelling was performed according to the aminoallyl labeling protocol. Briefly, cDNA was incubated at room temperature for 1 h with fluorolink monofunctional cy5 or cy3 (Amersham) followed by 15 min 4 M hydroxylamine treatment. Uncoupled dyes were removed using Qiaquick PCR purification columns (Qiagen) and mixed with 12 mg poly(dA) (Amersham), 60 mg yeast tRNA (Sigma) and 24 mg Cot-1 DNA (Invitroen). The probe was dissolved in 127 ml hybridization mixture containing 46% formamide, 9.5% dextran sulfate, 2x SSC and 0.2% SDS. The probe was heated to 70°C for 10 min and annealed at 37°C for 1 h. Slides were hybridized for 14 h at 37°C over-night (HyArray 12™, Perkin Elmer). After hybridization the slides were washed in the HybArray 12™ with 50% formamide, 2x SSC, pH 7 at 35°C for 15 min, followed b PI buffer (0.1 M sodium phosphate, 0.1% Igepal Ca630, pH 8) at room temperature and 3 washes of 0.2x SSC, 0.1x SSC and 0.01x SSC at room temperature followed by centrifugation. Arrays were scanned using a laser scanner (ScanArray Express, Perkin Elmer) and analysed using ImaGene version 5.6. Cy5 cy3 ratios are calculating by taking the log2 of the signal mean of each spot minus the median background mean of all spots on the array. This is followed by a standard normalization for spot intensity and calculation of the ratios. Keywords: other
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Contributor(s) |
van de Wiel MA, Costa JL, Smid K, Oudejans CB, Bergman AM, Meijer GA, Peters GJ, Ylstra B |
Citation(s) |
16288008 |
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Submission date |
Mar 01, 2005 |
Last update date |
Mar 16, 2012 |
Contact name |
Daoud Sie |
E-mail(s) |
d.sie@vumc.nl
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Phone |
+31 20 4442428
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Organization name |
Vrije Universiteit Medical Center
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Department |
Pathology
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Lab |
Microarray Core Facility
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Street address |
De Boelelaan 1117
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City |
Amsterdam |
ZIP/Postal code |
1081 HV |
Country |
Netherlands |
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Platforms (1) |
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Samples (5)
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GSM43959 |
Non resistant tumor x Gemcitabine resistant tumor |
GSM43960 |
Non resistant tumor (26A-4) x Gemcitabine resistant tumor (26G-3) |
GSM43962 |
Gemcitabine resistant tumor (26G-1) x Non resistant tumor (26A-2) |
GSM43963 |
Gemcitabine resistant tumor (26G-1) x Gemcitabine resistant tumor (26G-1) |
GSM43964 |
Cell line cA8 x Cell line cG7 |
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Relations |
BioProject |
PRJNA91645 |
Supplementary data files not provided |
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