52. RFLP mapping of the bacterial blight resistance genes Xa-3 and Xa-4

Satomi YOSHIMURA1,2,Rebecca NELSON1,Atsushi YOSHIMURA2,T.W.MEW1 and Nobuo IWATA2

1) International Rice Research Institute, P.O. Box 933, Manila, Philippines

2) Faculty of Agriculture, Kyushu University, Fukuoka, 812 Japan

Bacterial blight (BB), caused by Xanthomonas oryzae pv. oryzae (Xoo), is one of the most serious diseases of rice. The genetic basis of host resistance has been studied extensively, and 19 BB resistance genes have been identified from both cultivated rice and wild rice species. In this study, we determined the linkage relationships between two dominant resistance genes Xa-3 and Xa-4, and several RFLP markers previously mapped on the chromosome 11 of the rice genome by researchers in U.S.A. (McCouch and Tanksley 1991) and Japan (Saito et al. 1991). We established that the loci XNpb181 and XNpb78 are closely linked to the resistance genes.

To investigate the linkage relationships between these resistance genes and RFLP loci, we utilized near-isogenic lines for these resistance genes (IR-BB3 and IR-BB4), the donor cultivar for Xa-3 (Chugoku 45) and the recurrent parent used in the development of the near-isogenic lines (IR-24; Ogawa et al. 1988). Chugoku 45 (Xa-3), IR-BB3 (Xa-3) and IR-BB4 (Xa-4) were crossed with susceptible cultivar IR24. To examine the segregation of resistance and RFLP markers, F2 populations were inoculated with strain PXO61 (race 1) of Xoo and RFLP analysis was conducted on the same F2 plants. It has been previously reported that two RFLP loci on the chromosome 11 (XNpb181 = XNpb186 and XNpb78) showed polymorphisms between IR-BB3 and IR24, and between IR-BB4 and IR24 (Yoshimura et al. 1992). We used these RFLP markers for analysis of F2 populations derived from the 

Table 1.  Linkage analysis between Xa-3, Xa-4 and RFLP markers
===============================================================================
                                                                   Genetic
Gene pair     A-BB A-Bb A-bb aaBB aabb aabb1)   Total X2     Prob. map distance
                                                                    (cM)
==============================================================================
Xa-3-XNpb181     92       3      0       33     128  126.12   p<O.O1 2.3+/-1.34
Xa-3-XNpb78    19   70    4   0     1    34     128  134.43   p<O.O1 3.5+/-1.65
Xa-3-RG303     14   26    8   0     2    16      66  44.93    p<O.O1 14.9+/-4.64
Xa-3-CDO520    14   29    5   0     4    14      66  35.33    p<O.O1 13.7+/-4.46
Xa-3-CDO365    10   29    6   0     7     9      61  15.33    p<O.O1 23.4+/-5.90
Xa-3-RG2        9   31    8   0    12     6      66   10.73   0.1<p<.05 39.5+/-6.8
Xa-4-XNpb181   16   28    1   0     0    15      60  57.73    p<O.O1  1.7+/-1.68
Xa-4-XNpb78    15   29    1   0     0    15      60  57.53    p<O.O1  1.7+/-1.66
==============================================================================
1) Genotype of each F2 class is indicated. Capital letters A and B show the
alleles at two loci from resistant parent and small letters a and b from
susceptible parent IR24.  2) Calculated based on the ratio 9: 3: 3: 1 (df =
3).  3) Calculated based on the ratio 3: 6: 3: 1: 2: 1 (df 5).

Fig. 1. Gene arrangement of Xa-3(A) or Xa-4(B) and RFLP markers. The genetic distances are given in cMorgans.

crosses IR24/IRBB3 and IR24/IR-BB4. For RFLP analysis of Chugoku 45/IR24 cross, other RFLP markers on the chromosome 11 were also used (RG2, RG303, CDO365 and CDO520; McCouch and Tanksley 1991).

The map distances between resistance genes and RFLP loci are shown in Table 1. The data obtained from the Chugoku 45/IR24 and IR24/IR-BB3 F2 populations were combined to calculate the map distances between Xa-3, XNpb181 and XNpb78. Fig. 1 shows the chromosomal arrangement of the resistance genes and RFLP loci, integrating portions of the previously derived maps of the chromosome 11 (McCouch and Tanksley 1991, Saito et al. 1991). The results suggested that Xa-3, Xa-4, XNpb181 and XNpb78 fall outside the map published by McCouch and Tanksley (1991). Ogawa et al. (1986) suggested that the loci of Xa-3 and Xa-4 were allelic or tightly linked. Our filidings are consistent with their conclusions.

It is usuallay impossible to distinguish homozygosity of dominant genes based on phenotypic analysis. RFLP markers closely linked to such genes can be used to identify homozygous individuals, even without inoculation. We are currently using RFLP markers linked to BB resistance genes to select rice lines carrying multiple resistance genes. Closely linked molecular markers also provide a starting point for map-based cloning of resistance genes.

References

Ogawa, T., R. E. Tabien, G. A. Busto, G. S. Khush and T. W. Mew, 1986. The relationship between Xa-3, Xa-4 and Xa-4b for resistance to rice bacterial blight. Rice Genet. Newsl. 3: 83-84.

____, T. Yamamoto, G. S. Khush, T. W. Mew and H. Kaku, 1988. Nearisogenic lines as international differentials for resistance to bacterial blight of rice. RGN 5: 106-107.

Saito, A., M. Yano, N. Kishimoto, M. Nakagahra, A. Yoshimura, K. Saito, S. Kuhara, Y. Ukai, M. Kawase, T. Nagamine, S. Yoshimura, O. Ideta, R. Ohsawa, Y. Hayano, N. Iwata and M. Sugiura, 1991. Linkage map of restriction fragment length polymorphism loci in rice. Jpn. J. Breed. 41: 665-670.

McCouch, S.R. and S.D. Tanksley, 1991. Developnent and use of restriction fragment length polymorphism in rice breeding and genetics. Rice Biotechnology, p. 109-133. International Rice Research Institute, Manila, Philippines.

Yoshimura, S., A. Yoshimura, A. Saito, N. Kishimoto, M. Kawase, M. Yano, M. Nakagahra, T. Ogawa and N. Iwata, 1992. RFLP analysis of introgressed chromosomal segments in three near-isogenic lines of rice for bacterial blight resistance genes, Xa-1, Xa-3, and Xa-4. Jpn.J. Genet. 67: 29-37.