MOLECULAR DIFFERENTIATION AND MAP-BASED LOCALIZATION OF DISEASE RESISTANCE GENE ANALOGS IN HEXAPLOID OAT

Eastern Cereal and Oilseed Research Center, Agriculture and Agri-Food Canada, Ottawa, ON K1A 0C6 Canada

The sequences of several plant disease resistance genes and their encoded proteins show similar structural motifs in specific domains. In this study, 5 degenerate oligonucleotide primers were designed from the conserved coding domain sequences of tobacco mosaic virus resistance gene N, Arabidopsis bacterial blight resistance gene RPS2, flax rust resistance gene L6, and rice Xanthomonas resistance gene Cf-9, and were used to amplify related sequences from various cultivars of hexaploid oat which differed in resistance to different races of rust diseases. Alignment of 102 cloned DNA sequences from 11 cultivars revealed that at least 16 classes of resistance gene analogs (RGAs ) were detected. However, only 2 classes of RGAs in oat can be translated into intact proteins which contain a typic LRR domain. The other classes of RGAs contain short inserts or point mutations which interrupted the coding domain in the middle or near the 3' end of the sequences. Southern blot hybridization with translatable sequences ( eg. R4-1r ) showed a clear polymorphism among different cultivars. Genetic mapping of several of the translatable sequences of RGAs located them on linkage group 6 and 17 in a Kanota x Ogle mapping population. The location on the group 17 is near a mapped crown rust resistance gene locus. Mapping of other RGAs is in progress. Phylogenetic analysis and BLAST searches showed that most of the RGA sequences were similar to known RGAs from rice and barley and some of them are closely related to a cutinase gene, an infection response serine protein gene, insect resistance gene, or other known genes. Therefore, our data indicate that most biotic resistance genes and analogs share common process during plant evolution, the conserved domains in these genes can be used to design degenerate primers to amplify related resistance genes and analogs with a vast spectrum of resistance in crop plants.