1.2.2S
IDENTIFICATION AND SYNTENIC RELATIONSHIPS OF RESISTANCE GENE HOMOLOGUE LOCI IN CEREAL GENOMES

J KURTH, D LEISTER and P SCHULZE-LEFERT

The Sainsbury Laboratory, John Innes Centre, Colney Lane NR4 7UH, UK

Background and objectives
Plant resistance reactions to particular pathogens are triggered by corresponding resistance (R) genes in the host and avirulence (Avr) genes in the pathogen. Several dicot and one monocot R genes to diverse pathogens have been isolated so far and reveal structural similarities of the deduced proteins [1]. One class comprises RPS2, RPMI, N, L6, M, RPP5, Prf and 12C1. These genes contain both a 5' terminal nucleotide-binding site (NBS) and 3' terminal leucine-rich repeat (LRR) which is variable in length. A conserved stretch within the NBS domain and an adjacent 5'-located conserved peptide motif enabled a PCR-based appproach using degenerate primers to isolate R-like genes from dicot species [2]. The isolation of R genes from cereals with complex genomes is hampered by their large genome size. So far only one race-specific R gene, Xa-21 from rice has been isolated.

A fundamental feature of monocot species is their conserved gene order (synteny) [3]. This may facilitate map-based cloning from complex cereal genomes by using DNA marker information from syntenic intervals between simpler model grass species. Here we describe the isolation and the results of a comparative mapping of NB S-LRR homologues from rice and barley.

Results and conclusions
We have used conserved domains in the major class of dicot resistance genes (NBS-LRR) to isolate related gene fragments via PCR from the monocot species rice and barley. We show that this approach is feasible for monocot plant species with low- and high-complexity genomes. DNA of a low-complexity genome such as rice can be directly submitted to PCR with degenerate primers, whereas the isolation of NBS-LRR homologues from the highly complex barley genome was achieved by genomic subtraction or RT-PCR procedures before PCR. In total, 17 NBS-LRR homologues were isolated from rice and nine from barley.

Comparison of the deduced peptide sequence of dicot R genes and monocot R-like genes revealed shared motifs but provided no evidence for a monocot-specific signature. Mapping of these genes in rice and barley showed linkage to genetically characterized R genes and revealed the existence of clusters, each harbouring at least two highly dissimilar R-like genes. Diversity was detected intraspecifically with wide variation in copy number between varieties of a particular species. Interspecific analysis of R-like genes frequently revealed non-syntenic map locations between the cereal species rice, barley and foxtail millet, although tight co-linear gene order is a hallmark of monocot genomes. The observed data indicate a rapid evolution of R genes in related cereal species and suggest possible mechanisms to generate diversity in resistance loci.

References
1. Baker B, Zambryski P, Staskawicz B, Dinesh-Kumar SP, 1997. Science 276, 726-733.
2. Leister D, Ballvora A, Salamini F, Gebhardt C, 1996. Nature Genetics 14, 421-429.
3. Moore G, Devos KM, Wang Z, Gale MD, 1995. Current Biology 5, 737-739.