3.4.11
APPLICATION OF KNOWLEDGE ABOUT RESISTANCE GENE ORGANISATION IN ARABIDOPSIS THALIANA FOR GENETIC IMPROVEMENT OF BRASSICA OLERACEA

T WILKES1, D LECKIE1, N COGAN1, C RYDER1, S BREEDS1, P GORDON1, I PARKIN2, P BITTNER-EDDY1, K WILLIAMS3, J BEYNON1, I CRUTE1 and E HOLUB1

1Horticulture Research International, Wellesbourne, Warwick CV35 9EF, UK; 2Agriculture & AgriFood Canada, 107 Science Place, Saskatoon, Saskatchewan, Canada; 3Plant Research Centre, S. Aust. R & D Inst., Adelaide SA5001, Australia

Background and objectives
At least 45 wild-type genes (so-called R-genes) conferring resistance to viral, bacterial, fungal and oomycete parasites have been mapped throughout the Arabidopsis thaliana genome [1]. Twenty-five of these confer downy mildew resistance via pathotype-specific recognition of Peronospora parasitica (RPP), a biotrophic oomycete. In contrast, only a small number of disease resistance genes have been identified in Brassica species and few of these have been mapped.

Two approaches have been taken to utilise the information available from A. thaliana to facilitate the identification, mapping and cloning of resistance genes from Brassica oleracea. Firstly, an investigation of the degree of synteny between a Major RPP gene complex (MRC-F) on the bottom of chromosome 3 of A. thaliana and corresponding regions in B. oleracea was carried out. Specific DNA sequences from the RPP1 gene family from the MRC-F region were used as heterologous probes to identify homologues in B. oleracea. The second approach was to identify functional resistance genes in B. oleracea and generate associated AFLP markers which could be placed on an A. thaliana map to enable the identification of further markers and candidate homologous genes from A. thaliana.

Results and conclusions
Twelve RFLP markers which delimit the MRC-F region in A. thaliana were used to perform segregation analyses on thirty lines from a population of microspore derived doubled haploid lines produced from a highly polymorphic B. oleracea cross. Triplication of the B. oleracea genome was expected [2]. Four candidate regions, as defined by co-segregation of several polymorphic markers, are currently being examined: five markers co-segregate on linkage group 1 (LG1) of B. oleracea, three co-segregate on the top of LG3, nine co-segregate on the bottom of LG3, and three co-segregate on LG9. A BAC library generated from one of the parents of the B. oleracea mapping population has been screened using a member of the RPP1 gene family as a probe from the MRC-F region. Thirty-seven clones have been identified for further analysis.

Eight sources of resistance to P. parasitica and seven sources of resistance to Albugo candida (the causal agent of white blister) have been identified from a large-scale screening of 410 accessions of B. oleracea. Selected resistant plants have been crossed with a susceptible rapid-cycling line of B. oleracea and F2 populations produced. Segregation analyses have been carried out and all the resistant responses appear to be under simple genetic control. DNA from F2 plants has been used to produce resistant and susceptible bulks from each cross for AFLP analysis. Markers are being generated for four of the sources of disease resistance.

References
1. Holub EB, Beynon JL, 1996. Adv. Bot. Res. 24, 228-273.
2. Kowalski SP, Lan T-H, Feldmann KA, Paterson AH, 1994. Genetics 138, 499-510.