Project description:We used three different strains of Pseudomonas syringae pv tomato DC3000 to investigate systemic responses to infection in Arabidopsis and the development of SAR. Wildtype DC3000, the hrpA mutant and DC3000 carrying the avirulence gene avrRpm1 were syringe infiltrated into 4-week-old plants at a concentration of 10e8 cfu/ml. At least 5 leaves per plant were infiltrated and at least 10 plants were pooled for each sample. Systemic, uninfected tissue was then harvested at 8, 12 and 21h after inoculation. Three independent experiments were carried out to give three biological replicates for each timepoint. 27 samples were used in this experiment.
Project description:The sfr3-1 mutation causes freezing-sensitivity in Arabidopsis thaliana. The mutated gene has been identified by positional cloning and is currently being characterised. The mutant appears normal when grown in the warm (no phenotype has been identified associated with such growth). However, following cold acclimation and subsequent freezing mutant plants are severely damaged whilst wild type plants are not. This suggests that sfr3 is deficient in the cold acclimation process. Micro-array analysis will enable the identification of any transcriptional changes during the cold acclimation process. This information will then be used, together with information obtained by gene characterisation, in order to more fully understand the nature of the sfr3 mutation. 8 samples were used in this experiment.
Project description:Transcriptional comparison of Arabidopsis thaliana pvip1;pvip2 RNAi double mutants against Col-0 wildtype. Tissue used: whole rosettes of 4-week-old plants grown under short-day conditions (8h light; 20oC). 3 biological replicates for each. Control = Col-0 and mutant = pvip1;pvip2. PVIP = Potyvirus-VPg-interacting protein (Dunoyer et al. 2004 J. Virol. 78: 2301-2309). 6 samples were used in this experiment.
Project description:Reciprocal crosses of a diploid Arabidopsis with different ploidies results in different endosperm development patterns and phenotype. Typically if the ploidy is higher on the maternal side, there are fewer endosperm cells which cellularize early, and conversely, if the paternal ploidy is higher, there is more number of endosperm cells which cellularize late. Crosses involving diploid and tetraploid Arabidopsis (C24) are viable, whereas crosses involving the diploid and hexaploid, even though exhibit the above mentioned directional trend in endosperm development, abort (Scott et al 1998). The 'maternalised' and 'paternalised' development of endosperm is also observed in crosses involving some Arabidopsis mutants. Mutants in the fis class of genes, e.g. fis1-medea, when crossed with a diploid Arabidopsis (pollen parent) show endosperm development-seed development similar to a diploid (seed parent) crossed with hexaploid pollen parent. Reciprocal crosses of homozygous met1 and diploid Arabidopsis also exhibit reciprocal trends in endosperm development, where a homozygous met1 mutant (seed parent) crossed with diploid is similar in phenotype to a diploid (seed parent) - tetraploid cross. Endosperm development in the reciprocal cross has phenotypic similarity to the tetraploid (seed parent)-diploid cross (Adams et al 2000). We are interested in understanding gene profiles and trends in expression underlying the endosperm development in the interploidy crosses as well as the fis and met1 mutant. 11 samples were used in this experiment.
Project description:The aim of this project is to identify endogenous targets of yet unidentified epigenetic mutants. Mutants were discovered in a forward genetic approach using T-DNA insertion mutagenesis. Pools of 25 seedlings, grown in-vitro, will be used to examine changes in gene expression. 9 samples were used in this experiment.
Project description:Aim: To understand the role of DEK1 in Arabidopsis development. Background: DEK1 has an essential role during embryogenesis and is involved in the maintenance of the epidermis. Tissues: Comparison of wild-type and ADEK1 calpain-overexpressing line. 9 samples were used in this experiment.
Project description:Our aim is to identify circadian transcripts that are co-regulated with [Ca2+]cyt, with the eventual goal of identifying genetic regulators and targets for circadian oscillations of [Ca2+]cyt. We have identified two conditions in which [Ca2+]cyt behaves differently to other circadian outputs. 1. Treatment of plants with nicotinamide, a metabolic inhibitor of ADPR cyclase, abolishes the circadian oscillations of [Ca 2+]cyt. However, leaf movement, CCA1, LHY, TOC1 and CAB transcript abundance and CAB promoter activity are all rhythmic albeit with a longer period (Dodd et al., 2007). 2. The toc1-1 mutant, which shortens the circadian period of all other rhythms tested, has no effect on the period of [Ca2+]cyt oscillations (Xu et al., 2007). We will measure the circadian regulation of transcript abundance in wild type (C24), toc1-1 and nicotinamide (C24)-treated plants. Method: Wild type (C24) and toc1-1 seeds were sown on 1/2MS 0.8% agar plates and imbibed at 4 C for 48 hours. Seedlings were grown in LD cycles of 12hL:12hD at 19 C for 11 days to entrain the oscillator. Following transfer to LL at dawn on the 12th day, 50% of the wild type seedlings were dosed with 50 mM nicotinamide every 2 hours over the entire course of the experiment. Wild type, toc1-1 and nicotinamide-treated seedlings (approx. 100 for each sample, excluding roots) were harvested at 4 hour intervals from 49 to 93 hours in LL (12 time points covering the entire third and fourth circadian cycles). Two independent replicates of the whole experiment will be hybridised. 72 samples were used in this experiment.
Project description:It has been established that the frequently reported early flowering phenotype of the phyB mutant is very sensitive to small changes in ambient temperature (Halliday et al., 2003). A 6oC drop in temperature to 16oC is sufficient to abolish the pronounced early flowering phyB phenotype observed at 22oC. Using Affymetrix technology we will identify genes that are differentially regulated at 16oC and 22oC in a phyB-specific manner using phyB mutant and wild type seedlings. 16 samples were used in this experiment.
Project description:MALE STERILITY 35 (MS35/ MYB26) is a R2R3-type MYB transcription factor (Steiner-Large et al., 2003). MS35 is suggested to be a key regulator in the process of lignified secondary thickening in the anther endothecium which provides mechanical force for anther dehiscence (Dawson et al., 1999). The ms35 mutant bears non-dehiscent anthers and thus exhibits a male sterile phenotype. This microarray experiment aims to identify genes controlled by MS35 and thus establish the precise role MS35 plays during anther development. We have demonstrated that MS35 is expressed in the anther during pollen mitotic divisions. Comparisons of gene expression will be carried out between anthers from the Ler.gl (wild-type control) and ms35.gl (mutant) to identify genes with altered expression due to the ms35 mutation. The use of the gl marker will enable a prompt identification of the ms35 mutant amongst the segregating population. The wild-type control line was chosen to be Ler.gl which ensures that the changes of gene expression are not due to the gl mutation. Plants were grown in growth chambers at 22h of daylight (167 umol.m2.sec-1) at 24 C and 2h of night at 24 C. Developmental stages of anthers were defined by bud position and DAPI staining. As soon as the plants showed three fertile or sterile siliques, anther and filaments were collected from both lines at Pollen Mitosis I (PM I), bicellular, and PM II stage, separately. RNA was extracted using RNeasy Mini Kit (Qiagen, USA) and digested with DNase to eliminate genomic DNA contamination. The identified genes will be further analysed using bioinformatics and genomic (knockout) resources that are available for Arabidopsis. 12 samples were used in this experiment.
Project description:Brassinosteroids (BRs) are steroid hormones that are essential for the development of plants. A tight control of BR homeostasis is vital for modulating their impact on various growth responses. However, whilst it is recognized that the rapid adaptation of de novo synthesis plays a key role in adjusting required BR levels, our knowledge of the mechanisms governing feedback control is limited. We identified the transcription factor CESTA as a novel regulator of BR biosynthesis. ces-D was isolated in a screen of Arabidopsis mutants by BR over-accumulation phenotypes. Loss-of-function analysis and the use of a dominant repressor version revealed functional overlap among CESTA and its homologues and confirmed the role of these basic helix-loop-helix proteins in the positive control of BR biosynthetic gene expression. In this experiment we compare the transcriptome of the CESTA mutant to wild-type plants using the Affymetrix ATH1 array. 4 samples were used in this experiment.