Transcription profiling of Arabidopsis Col-0 plants sprayed with sodium nitroprusside (SNP)
ABSTRACT: 22 day old Arabidopsis Col-0 plants were sprayed with 0.5 mM sodium nitroprusside (SNP), corresponding controls were sprayed with water, and leaves harvested 3 hours after spraying and frozen in liquid nitrogen.
Project description:We are studying the tree P. euphratica growing in its natural habitat in the Negev desert in Israel. We have used leaf RNA samples from trees growing from four different areas in the desert valley Ein Avdat with contrasting growth conditions, with the primary factor being how much water the trees have access to. Area A trees grow close to a stream. Area B trees are further away from the stream. Area C trees are growing on a slope with no water. Parking place trees grow at a parking place (1km from Area A, B and C) where there is a water irrigation system, and these trees are regularly watered once a week. The control sample in each hybridization is always RNA isolated from a pool of 5 trees from the parking place. The hybridizations are comparing the following: Area A -- parking place Area B -- parking place Area C -- parking place Each hybridization is done with a dye swap and three different biological repeats for a total of 18 hybridizations.
Project description:Rooted plantlets of Populus euphratica were transferred to aerated hydroculture with Long Ashton nutrient solution, grown for 3.5 months at 22°C, 150 µmol m-2 s-1 photosynthetically active radiation with a photoperiod of 16h light. After reaching an average height of 0.83 m, the media was supplemented with 150 mM NaCl (Salt-treated) or maintained under control conditions (control). During the experimental phase, light was continuously supplied to avoid interference of light/dark transitions. <br>Roots and leaves of six plants were harvested separately after 3h, 6h, 12h, and 24h of salt exposure and of controls at the corresponding time points (= 48 plants per experiment). The material of each harvest of 6 plants was pooled, yielding one leaf and one root sample of controls and one leaf and one root sample of salt-treated plants, respectively (=16 samples per experiment).<br>The experiment was performed three times yielding three biological replications (=48 samples in total).<br>After extraction of RNA and cDNA synthesis, each extract was labelled with Cy3 and Cy5, respectively (yielding 96 labelled extracts). Extracts from e.g. 3h-salt-treated leaf exp. 1 (cy3) was hybridized with 3h-control leaf exp.1 (cy5) with microarrays (UHPB-P.euphratica-10K-5; A-MEXP-93)containing unique 7,662 ESTs of P. euphratica from stress-induced cDNA libraries (Brosche et al., Genome Biology 2005, 6:R101). Dye swap was perfomed, thus usually yielding 6 arrays per timepoint and plant tissue (3 biological, 3 technical replicates). In some cases hydridizations were not successfull. Therefore, for roots harvested at timepoints 12h and 24 h only 4 arrays per timepoint were used. For roots harvested after 3h an additional mixed sample of all 3 biological replicates was performed. One leaf array at 6 h was not sucessfull. Therefore, residual labelled extract from salt treated plants of exp1 was hydridized with controls at 6h from exp.2. <br>
Project description:Plantlets of P. euphratica (clone B2) were obtained by in vitro culture. After ex vitro acclimation, they were transferred and acclimated to greenhouse conditions and transplanted into 7.5L-pot filled with peat-sand mix (50/50 V/V) and were pruned (following some leaf shedding due to transplantation shock). In the greenhouse, climate conditions depended on outside weather but temperature was regulated in order to remain between 15 and 30°C. A moderate, increasing drought stress was applied and controlled for 6 weeks. Soil volumetric water content was measured one to two times a day depending on the intensity of the stress (TDR and weighting). Plantlets were progressively brought to 5 stress levels [soil volumetric water content]: 10% (harvest 1, 11 days from start of experiment), 7.5% (harvest 2, 16 days from start of experiment), 5% (harvest 3, 22 days from start of experiment), 4% (harvest 4, 26-32 days from start of experiment) and back to fully available water (harvest 5, 10 days after fully watering of the trees; 36 days from start of experiment). Leaves and roots (including control and stressed trees) were harvested for each stress level. At harvest 1-3, samples from five trees were harvested. At harvest 4, samples from two trees were harvested at each of day 26, 29 and 32. At harvest 5, samples were harvested from two control trees and four re-watered trees. <br><br> For the array experiment: leaf or root samples were pooled before RNA isolation to give three repeats by dividing the five samples 2+2+1. For harvest 4; samples were pooled from each of the two trees sampled at day 26, 29 and 32 to give three biological repeats. In harvest 5 samples were pooled 2+2 to give two biological repeats. Each array experiment was done with a dye swap to give 28 hybridizations for the leaf and 28 hybridizations for the root experiment.
Project description:Control (Col-0) and mutants (rcd1-1, rcd1-3, rcd1-4 and sro1-1) were grown for 22 days under control conditions. Whole rosettas were harvested and analyzed for changes in gene expression between Col-0 and the mutants.
Project description:23 day old Arabidopsis plants, wildtype Col-0 and mutant rcd1, were compared to determine which genes are over- or under expressed in the mutant. Six biological repeats were pooled in pairs of two for labellings. All labellings/hybridizations were done with a dye swap technical repeat (e.g. in total six labellings were used). The Turku Arabidopsis 8K arrays were used for hybridizations. For the hybridizations the samples were split in two halves with each half hybridized to Turku Arabidopsis 8K slide 1 and Turku Arabidopsis 8K slide 2 (e.g. in total 12 hybridizations were performed).
Project description:We have used transgenic ethylene-insensitive birches (Betula pendula), which express the Arabidopsis ethylene receptor gene ETR1 carrying the dominant mutation etr1-1, to investigate the role of ethylene in short day (SD) -induced responses in the shoot apical meristem in birch. Wild-type birch (clone V5834) and two ethylene-insensitive lines in this background (BPetr1-1-35 and BPetr1-1-86; see Plant Physiol 132: 185-195) were exposed to SD. After 12, 16 and 20 days under SD, apices of branches of three trees were pooled before RNA extraction from each sample. To study the ethylene-dependent SD-transcriptome in birch apices, the RNA extracts of lines BPetr1-1-35 and BPetr1-1-86 were separately compared with the reference, wild-type V5834, at the three time points (12SD, 16SD, 20SD) resulting in altogether six microarray hybridizations.
Project description:Gene content in various Enterococcus faecalis strains compared to E. faecalis V583. Strains have been compared to the V583 strain by comparative genomic hybridization using genome-wide PCR-based microarrays representing the V583 genome. Genes have been deemed "present" or "divergent" in the various strains.