Arabidopsis thaliana col-0, inoculated with Erysiphe cichoracearum, with Si supplementation
Extracted molecule
total RNA
Label
Cy5
Hybridization protocol
Hybridization was performed following Agilent's oligonucleotide microarray hybridization user's manual and Agilent's in situ Hybridization Plus kit. A volume of 200 µL of combined Cy3- and Cy5-labeled cDNA targets were denatured at 98 °C for 3 min and cooled to room temperature. They were mixed with 50 µL of 10x control targets, followed by the addition of 250 µL of 2x hybridization buffer. The 500 µL of reaction mix was applied to each Agilent 44K Arabidopsis 3 microarray (37,537 features), and hybridized in a hybridization rotation oven at 60 °C for 17 h. The slides were disassembled in 6 x SSC, 0.005% Triton X-102, washed first with 6xSSC, 0.005% Triton X-102 for 10 min at room temperature, then with 0.1 xSSC, 0.005% Triton X-102 for 5 min on ice, and dried using a nitrogen-filled air gun.
Scan protocol
The arrays were scanned using a dual-laser DNA microarray scanner (Agilent). The data was then extracted from images with Agilent’s Feature Extraction software 7.5.
Description
Supplementary file contains data obtained after scan
Data processing
The R software 2.3 and the LIMMA package were used to normalize the microarray data (loess method for within array normalization followed by quantile method for between arrays normalization)
