43. 
Comparison of RGA and RFLP markers for determining the genetic relationship in rice
 
J.Y. ZHUANG’, J.L. WU’, Y.Y. FAN’, H. LEUNG2 and K.L. ZHENG’
1) 
Biotechnology Department, China National Rice Research Institute, Hangzhou, 310006, China
2) 
Entomology and Plant Pathology Division, International Rice Research Institute, 1099 Manila, Philippines
 

 
     It has been demonstrated that resistance gene analogs (RGAs) detected using denatured polyacrymide gel electrophoresis can reveal high polymorphism in rice, offering an opportunity of using RGA markers for a rapid germplasm identification in rice. However, the feasibility of using RGA markers for germplasm studies would depend greatly on whether the results revealed by RGA markers can represent the genetic relatedness over the entire rice genome.
     In our study, a common set of 16 indica rice varieties was used for RFLP and RGA analyses. To infer the genetic relationships among the varieties, each polymorphic band was scored as “1” for presence and “0” for absence. Genetic similarity between two varieties was computed as the number of common bands divided by the total number of bands in two varieties (Nei 1987, formula 5.53). The frequency of polymorphism, computed as the number of polymorphic variety pairs divided by the total number of variety pairs, was used to compare the polymorphism detecting ability of polymorphic RGA and RFLP bands.
     Ninety-seven RFLP probes were selected to cover the 12 rice chromosomes based on published mapping information. In combination with restriction endonuclease EcoR I, 39 of the 97 probes detected polymorphism among the 16 varieties and produced a total of
90 polymorphic fragments. The average number of polymorphic bands detected by each of the 97 probes was 0.93 only, and that over the 39 polymorphic probes was 2.3. All the varieties can be distinguished, and the minimum number of polymorphic bands between any two varieties was 4. Over the 90 polymorphic bands, the average frequency of polymorphism between all pairs of varieties was 0.299.
     Four pairs of RGA primers were used and generated 111 polymorphic bands among the 16 varieties. All the varieties can be distinguished, and the minimum number of polymorphic bands between any two varieties was 11. The average number of polymorphic bands detected by one pair of RGA primer was 27.75. Over the 111 polymorphic bands, the average frequency of polymorphism between all variety pairs was 0.376. In addition, the number of polymorphic bands generated by different primer pairs ranged as 28—34, and the average frequency of polymorphism ranged as 0.365—0.395. This indicated that different primer pairs have similar polymorphism detecting ability. Although the distribution of all RGA markers was unknown and some RGA markers may be located as clusters, we could expect that a small number of RGA primers would produce sufficient information for germplasm evaluation in rice because of the great ability of RGAs on detecting polymorphism.
     The value of similarities between all pairs of varieties were computed based on the 90 polymorphic RFLP fragments and the 111 RGA bands, respectively. The similarity coefficients generated by RGA markers were plotted against those generated by RFLP markers (Fig. 1). Highly significant correlation (r=0.73) was obtained, indicating that the cultivar relatedness revealed by four pairs of RGA primers was comparable to that revealed by RFLP markers distributed over a large proportion of rice genome. In addition, the genetic similarities revealed by RGA markers were generally lower than that revealed by 
RFLP markers (Fig. 1), indicating that RGA might be more efficient for detecting polymorphism.

References

Nei, M., 1987. Molecular evolutionary genetics. Columbia University Press, New York, pplO6.