Project description:AtGenExpress: A multinational coordinated effort to uncover the transcriptome of the multicellular model organism Arabidopsis thaliana (Hybridisations done at NASC) The activity of genes and their encoded products can be regulated in several ways, but transcription is the primary level, since all other modes of regulation (RNA splicing, RNA and protein stability, etc.) are dependent on a gene being transcribed in the first place. The importance of transcriptional regulation has been underscored by the recent flood of global expression analyses, which have confirmed that transcriptional co-regulation of genes that act together is the norm, not the exception. Moreover, many studies suggest that evolutionary change is driven in large part by modifications of transcriptional programs. An essential first step toward deciphering the transcriptional code is to determine the expression pattern of all genes. With this goal in mind, an international effort to develop a gene expression atlas of Arabidopsis has been underway since fall 2003. This project, dubbed AtGenExpress, is funded by the DFG, and will provide the Arabidopsis community with access to a large set of Affymetrix microarray data. As part of this collaboration, we have generated expression data from 80 biologicaly different samples in triplicate. Series 2: Base line experiment for pathogen infection. Growth and infection conditions: Seeds will be stratified for 3 days at 4°C and sown on soil. Plants are grown at 22°C under a 8/16 hour light/dark regime in Percival growth chambers. All pathogen treatments will be performed on leaves of 5-weeks old plants. Bacterial infiltrations will be performed with 10-8 cfu/ml in 10 mM MgCl2. 5x105 spores of Phytophthora infestans in water will be applied to leaf surfaces. LPS (100 µg/ml), HrpZ (1 µM), NPP1 (2 µM) or flagellin (1 µM flg22) will be applied in water. Experiments should be performed in triplicate at the time points indicated below. Experimenter name = Dierk Scheel , Frederic Brunner , Lore Westphal Experimenter institute = AtGenExpress Keywords: genetic_modification_design; organism_part_comparison_design; development_or_differentiation_design
Project description:Late blight of tomato is caused by the oomycete pathogen Phytophthora infestans. In our previous work, we identified and characterized a miR1918 in P. infestans (pi-miR1918), and showed that its sequence is similar to the sequence of tomato miR1918 (sly-miR1918). In this study, we used Arabidopsis thaliana pre-miR159a as a backbone to synthesize pi-miR1918 via PCR and mutagenesis. The artificial pi-miR1918 was used to investigate the role of miR1918 in tomato-P. infestans interaction. Trangenic tomato plants that overexpressed the artificial pi-miR1918 displayed more serious disease symptoms than wild-type tomato plants after infection with P. infestans, as shown by increased number of necrotic cells, lesion sizes and number of sporangia per leaf. The target genes of pi-miR1918 and sly-miR1918 were also predicted for tomato and P. infestans, respectively. qPCR analysis of these targets also performed during tomato-P. infestans interaction. The expression of target gene, RING finger were negatively correlated with miR1918 in the all Lines of transgenic tomato plants. In addition, we used the 5' RACE to determine the cleavage site of miR1918 to RING finger. These results suggested that miR1918 might be involved in the silencing of target genes, thereby enhancing the susceptibility of tomato to P. infestans infection.