In hybrid breeding programs involving cytoplasmic male sterility systems, restorers are routinely identified by testcrossing prospective lines with available CMS lines and evaluating F₁s for pollen and spikelet fertility. In IRRI's hybrid rice breeding program, hundreds of testcrosses are made and evaluated every season and lines showing >-80% pollen and spikelet fertility are designated as restorers. Using this protocol, up to 40% restorer frequency has been observed among elite indica lines. The majority of the elite lines are found as partial or incomplete restorers which cannot be utilized as parents to make experimental rice hybrids.

Fertility restoration for 'WA' cytoplasm has been found to be controlled by two Rf genes, one of which is stronger than the other (Govinda Raj and Virmani, 1988; Bharaj, et al., 1991). Chromosomal locations for such genes have been reported on chromosomes 7 and 10 (Bharaj et al., 1995), chromosome 1 (Zhang et al., 1997), four QTLs on chromosomes 2, 3, 4, and 5 (Zhu et al., 1996), two QTLs on chromosome 10 (Tan, 1998) and four QTLs on chromosomes 1, 7, 10 and 11 (Zhuang and Zheng, 2000). Govinda Raj and Virmani (1988) also provided evidence to indicate the presence of different sets of Rf genes in different restorer lines. Perhaps, complex inheritance and involvement of more than one gene discouraged hybrid rice breeders to use marker aided selection for fertility restoration.

We targeted the major 'WA' restorer gene located on chromosome 1 previously tagged with the PCR marker RG140STS. Polymorphism between restorer and non-restorer is achieved after digestion with PvuII (Figure 1). A total of 227 lines from the Source Nursery (2000 wet season) were analyzed by PCR and F₁s from testcrosses with IR68897A and IR70369A (with 'WA' cytoplasm) were grown and evaluated for pollen and spikelet fertility in the Testcross

Nursery (2001 dry season) at IRRI. Of the 126 lines identified by PCR as putative restorers, 105 (83%) exhibited restorer ability with one or both testers (Table 1). Among the 101 lines identified as putative non-restorers, only 12 (12%) were found to be restorers by testcross method.

Considering the fact that we targeted only one of the two genes which control restoration ability for 'WA' cytoplasm in rice, these results are very encouraging. They demonstrated for the first time the use of molecular markers for selecting restorers lines. Indirectly, these results also confirmed that the Rf gene targeted for marker aided selection in this study was a strong one. With the tagging of other Rf genes or QTLs using appropriate molecular markers, the efficiency of marker aided selection for restorer genes can be further improved. It should then be possible to routinely use molecular markers to select putative restorers of 'WA' cytoplasm among a set of elite lines. These putative restorers can then be testcrossed with appropriate CMS lines to confirm their fertility restoration and check the magnitude of heterosis. This

approach should considerably reduce the amount of work done in making and evaluating testcrosses to identify effective restorer lines in a hybrid rice breeding program.

References

Bharaj, T.S., S.S. Bains, G.S. Sidhu and M.R. Gagneja. 1991. Genetics of fertility restoration of 'wild abortive' cytoplasmic male sterility in rice, Oryza sativa L. Euphytica 56: 199-203.

Bharaj, T.S., S.S. Virmani and G.S. Khush. 1995. Chromosomal location of fertility restoring gene for 'wild abortive' cytoplasmic male sterility using primary trisomics in rice. Euphytica 83: 169-173.

Govinda Raj, K. and S.S. Virmani. 1988. Genetics of fertility restoration of WA type cytoplasmic male sterility in rice. Crop Sci. 28:787-792.

Tan, X.L., A. Vanavichit, S. Amornsilpa and S. Trangoonrung. 1998. Genetic analysis of rice CMS-WA fertility restoration based on QTL mapping. Theor. Appl. Genet. 96: 994-999.

Zhang, G., T.S. Bharaj, Y. Lu, S.S. Virmani and N. Huang. 1997. Mapping the Rf-3 nuclear fertility-restoring gene for WA cytoplasmic male sterility in rice using RAPD and RFLP markers. Theor. Appl. Genet. 94: 27-33.

Zhu, L., C. Lu, L. Shen, Y. Xu, P. He and Y. Chen. 1996. Using doubled haploid populations of rice for quantitative trait locus mapping. In: Khush, G.S. (ed) Rice Genetics III. Proc 3^rd Int Rice Genet Symp. IRRI, Manila, pp 631-635.

Zhuang, J.Y. and K.L. Zheng. 2000. Mapping major and minor QTL for rice CMS-WA fertility restoration. Rice Genet. Newsl. 17: 56-59.