Growth Conditions and Treatments: Arabidopsis thaliana seeds were surface sterilized. Approximately 15000 seeds(300 milligrams) were placed in a 1 L erlenmeyer flasks containing 300 mL of liquid Murashige and Skoog medium (Sigma, St Louis, MO, USA) containing 1.5% sucrose. These flasks were incubated with gentle shaking for 5 days under constant illumination at 22ºC. After this time, 15 ul of 80% ethanol was added as the mock control (BL treated received 15 ul of 2uM Brassinolide (BL) dissolved in 80% ethanol. The BL treatment results were not used and are thus not reported). Flasks were incubated for 3 hours with the ethanol. Whole seedlings from each treatment were collected, the medium was drained, and the seedlings were rapidly forzen in liquid nitrogen. The experiment was done alongside two other genetic backgrounds (wildtype and bzr1-1D) and was repeated three times (experiments 1-3) during different weeks. RNA and Microarray Methods: Total RNA was extracted from the plants using a modified Trizol method (Chomczynski and Sashi, 1987)(see also Protocols Manual at the AFGC web page http://www.arabidopsis.org/info/2010_projects/comp_proj/AFGC/RevisedAFGC/AFGC_Protocols_Dec_2001L.pdf) and purified with a silica membrane column (Qiagen, RNeasy). Fifteen micrograms biotinylated complementary RNA (cRNA) was prepared and used to hybridize ATH1 Arabidopsis GeneChips (Affymetrix) using the manufacturer’s protocols. The array images were analyzed with the Affymetrix GeneChip Operating Software (GCOS) 1.1 with the target intensity set to 500. These data were imported into GeneSpring 7.0. To remove chip-to-chip and experiment-to-experiment signal variation each array was normalized by the intensity values of the top 50th percentile of genes on this array. The individual detection values were then normalized to the corresponding detection values of the wildtype (Colombia-0) control for each experiment. The normalized values for the data set are reported along with the intensity values for this array. Keywords = Arabidopsis, hormone, brassinosteroid, det2, bzr1-1D Lot batch = 510690
Brassinosteroid mutant (bzr1-1D and det2) analysis
Data table header descriptions
ID_REF
As defined by Affymetrix, there are the probe set identifiers, each of which is unique to a specific probe set defining a specific reagion of a singal gene or set.
VALUE
This is the final calculated measurement for each probe set idendifier that has been made comparable across all samples and rows.
ABS_CALL
A qualitative measurement indicating if the probe set is detected (Present; P), not detected (Absent; A), or marginally detected (Marginal;M)
DETECTION P-VALUE
A p-value indicating the significance of the Detection call. A Detection p-value measures the probability that the discrimination scores of all probe pairs in the probe set are above a certain level, and that the target is likely to be Present.
Normalized Value of Control
A normalized measurement of control arrays for each probe set calculated in GeneSpring 4.2.
