An integrated informatics and resources platform for Reverse Genetics in dicots (RevGenUK)

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The genome sequences of Arabidopsis and rice have been completed and those of other species including Lotus japonicus, Medicago truncatula and Brassica rapa are currently underway. To uncover the function of the thousands of genes that will be discovered by these initiatives will be a major challenge. Reverse genetics uses genome sequence to permit the recovery of mutants in target genes for use in functional genomics. This is achieved in Arabidopsis using TILLING to obtain an allelic series of mutants and T-DNA insertion to generate null mutations. The latter uses the efficient transformation system available for this plant, but lacking in our target species, so we need to find alternatives. One such system, named de-TILLING, is based on deletions induced by fast-neutron (FN) mutagenesis. The use of both TILLING and de-TILLING mutation detection systems in a species permit the isolation of an allelic series of mutants including nulls.
A third method for studying allelic differences in genes is by using ecotype variation (Eco-TILLING). We have assembled or are in the process of assembling populations for TILLING in L. japonicus, M. truncatula and B. rapa, and a population of FN plants for M. truncatula. Mutation detection is usually carried out using gel-based systems that are inefficient and costly. We have developed an ABI3730 system that is highly efficient. We now wish to gather our existing populations, generate complementary populations and migrate all three mutation detections systems onto the ABI platform for our target species to develop a reverse genetics service for the community. To do this we propose to establish FN populations for L. japonicus and B. rapa, Eco-TILLING populations for all our target species and collate the data in an open-source database available over the web. Under the umbrella of the single technology platform, we will create a unique resource for mutation discovery and functional genomics for the benefit of research community.

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