Molecular tools for studying the Arabidopsis transcription factor ORFeome

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Genomic approaches have generated large Arabidopsis open reading frame (ORF) collections. However, tools are required to functionally characterise this ORFeome. In here we propose the development of two technology platforms based on an existing transcription factor (TF) ORFeome of 1200 full-length cDNAs: (1) We have developed a novel batch procedure to simultaneously recombine GATEWAY®-tagged cDNAs into a plant expression vector. This pool has been applied to build up an Arabidopsis thaliana TF ORF over expressing (AtTORF-Ex) seed stock library harbouring a near-complete collection of 115 AP2/ERFs TF. As part of this project we attempt to enlarge this library up to 1000 transgenic lines covering around two thirds of the Arabidopsis TF ORFeome. This resource will be used to screen for TFs mediating enhanced stress tolerance. (2) A high-throughput microtiter plate based protoplast transfection system has been set up which will be used to analyse the trans activation properties of the TF ORFeome (Protoplast Trans Activation System: PTA). By providing plasmid pools, those TFs can be rapidly identified, which are regulating a particular promoter. These molecular tools will significantly improve functional studies on TFs performed by the AFGN consortium and the Arabidopsis community.