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Status |
Public on Jan 26, 2011 |
Title |
PH1_tebuconazole treated_rep3 |
Sample type |
RNA |
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Source name |
PH1_complete medium_5 mg/l tebuconazole_12h
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Organism |
Fusarium graminearum |
Characteristics |
strain: PH1 (syn. NRRL31084)
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Treatment protocol |
After 24 h, CM medium of the treatment cultures was adjusted to 5 mg/l tebuconazole and incubation of all cultures continued for further 12 h.
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Growth protocol |
106 conidia were inoculated into 300 ml flasks each containing 100 ml complete medium (Leslie & Summerell, The Fusarium Laboratory Manual, Blackwell Publishing) and cultivated at 23°C with shaking at 150 rpm.
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Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was isolated from mycelia grown in vitro with the Qiagen RNeasy Plant Mini Kit following the manufacturer’s protocol. The optional on-column DNase digestion step was performed using RNase-free DNase (Qiagen).
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Label |
Cy3
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Label protocol |
For sample preparation and array processing the Agilent protocol “One-color microarray-based gene expression analysis (Quick Amp Labelling)” was used (http://www.chem.agilent.com).Briefly, the recommended volume of control RNAs (Agilent One-Color RNA Spike-In Kit) was added to 200 ng of total RNA. Thereafter, cyanine 3´- labelled cRNA was produced, using the Agilent Quick Amp Kit (one-color) according to the manufacturer’s protocol. Labelled cRNA was purified with the RNeasy mini kit (Qiagen). Yield and Cy3 incorporation rates were analyzed using a NanoDrop ND-1000 UV-VIS spectrophotometer. In addition, size distribution of cRNAs was assessed by a BioAnalyzer 2100.
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Hybridization protocol |
For hybridizations, 600 ng of Cy3-labelled cRNA was used. Hybridization was set up in an Agilent Microarray Hybridization Chamber (G2534A) and incubated in an Agilent Hybridization Oven (G2545A) at 65°C and 10 rpm for 17 h. Thereafter, suggested washing steps were performed.
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Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2565BA) using one color scan setting for 8x15K array slides (scan resolution: 5 µm resolution).
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Description |
Gene expression after 12 h - treatment with 5mg/l tebuconazole
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Data processing |
The microarray scan data were analyzed by Agilent's Feature Extraction software_v10.5 obtaining background subtracted and spatially detrended signal intensities. These 'gProcessedSignal' data were converted to gene signals by a median polish algorithm as implemented in the RMA algorithm. Gene signals were normalized between experiments by quantile normalization.
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Submission date |
Nov 03, 2010 |
Last update date |
Jan 26, 2011 |
Contact name |
Rayko Becher |
E-mail(s) |
rayko.becher@landw.uni-halle.de
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Organization name |
Martin-Luther-University Halle-Wittenberg, Institute of Agricultural and Nutritional Sciences
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Department |
Department of Phytopathology and Plant Protection
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Street address |
Betty-Heimann-Str. 3
|
City |
Halle/Saale |
ZIP/Postal code |
06120 |
Country |
Germany |
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Platform ID |
GPL11158 |
Series (1) |
GSE25114 |
Genome-wide expression profiling of Fusarium graminearum in response to azole fungicide treatment |
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