3.4.66
SATURATION MAPPING OF CONSERVED MOTIFS OF DISEASE RESISTANCE GENES IN RICE USING A DOUBLED HAPLOID POPULATION AND NEAR-ISOGENIC LINES

H LEUNG, MLC GEORGE, MR BARAOIDAN, MD VILLAMAYOR and AM MILLENA

Division of Entomology and Plant Pathology, International Rice Research Institute, Los Banos, Philippines

Background and objectives
Disease resistance genes from diverse plant species have conserved motifs that can be used to design PCR primers to amplify candidate resistance genes. High resolution electrophoresis and silver staining can be used to detect a large number of conserved motifs in the genomes of wheat, barley and rice [1]. We applied this method to rapidly map conserved motifs in rice using a doubled haploid (DH) population derived from IR64 x Azucena and segregation populations derived from near-isogenic lines (NILS) carrying single disease resistance genes.

Results and conclusions
Nineteen primer pairs corresponding to the conserved regions of leucine rich repeats (LRR), nucleotide binding sites (NBS), and protein kinases in several cloned resistance genes were used to amplify polymorphic DNA fragments in IR64 and Azucena. An average nine polymorphic markers per primer was detected between the parents. Approximately 120 markers have been placed onto a frame map of rice. While these markers are found on all 12 rice chromosomes, their distributions within a chromosome appear to be clustered. Twenty-seven (23%) of these markers fall into chromosomal regions containing major blast (Pi) and bacterial blight (Xa) resistance genes: Pi10 on chromosome 5; Pi2 and Pig on chromosome 6; Pi11 on chromosome 8; Pi7, Pi1, Xa3, and Xa4 on chromosome 11; and Pi6 and Pi4 on chromosome 12. No conserved motifs have yet been mapped perfectly to the resistance genes segregating in this DH population. Interestingly, some LRR and kinase conserved motifs map at the vicinity of putative QTL for resistance against blast. To tag specific resistance genes, we surveyed variations in the conserved motifs in NILs carrying single Pi and Xa genes. Considerable polymorphism was observed between the NILS, each with unique bands distinct from the recurrent parents. Within the Pi NIL series (in Co39 background), approximately 15% dissimilarity was detected [1]. For the Xa NIL series (in IR24 background), approximately 6% dissimilarity between lines was observed. We have generated F2 populations (200-400 individuals per family) segregating for all available Pi and Xa genes in the NILS. Co-segregation analysis between the agronomically important genes (Pi1, Pi2, Xa4, and Xa7) and conserved motifs is underway. The results gathered thus far demonstrate the efficiency of the genome scanning approach in saturating the rice genome with conserved motifs. Although the majority of the amplified DNA fragments are not necessarily functional resistance genes, they may represent genes involved in other signal transduction pathways. Thus, construction of a rice map saturated with these motifs will be helpful for identification of genes involved in response to other biotic and abiotic stresses.

References
1. Chen XM, Line RF, Leung H, 1998. Theor. Appl. Genet. (in press).