31. Identification of RAPD markers linked to Se-3(t), a gene enhancing the level of
    photoperiod sensitivity in rice

    M. Maheswaran 1,3, D. J. Mackill2, Ning Huang 1, S. R. Sreerangasamy3 and S. R. McCouch4
        1. Genome Mapping Laboratory, PBGB, IRRI
        2. USDA-ARS, Dept of Agronomy & Range Science, University of California, USA
        3. Tamilnadu Agricultural University, Coimbatore, India
        4. Department of Plant Breeding and Biometry, Cornell University, lthaca, USA

    The genetic analysis done on different cross combinations of photopeirod insensitive and sensitive rice varieties revealed the presence of two genes viz, Se-I and .Se-3(t) controlling photopeiod sensitive phase and critical day length respectively. Kinoshita (1986) reported that photoperiod sensitivity gene Se-I is closely linked with isozyme marker Pgi-2. Based on this report Pgi-2 alleles were used to determine if any of the genes conferring photoperiod sensitivity were allelic to Se-I. Results indicated that Puang Rai 2, Sac Nau, and Nam Saugi 19 all had a dominant gene conferring sensitivity at the Se-I locus, (Poonyarit et al., 1989). The details of the parents and progenies involved in
 
 
 

Table 1. The parents involved in gene tagging for photoperiod sensitivity
Parent Origin Duration Genotype Phenotype
IR 26760-27 IRRI 145 se lse 1 Se 3 Se 3 weakly sensitive
Nam Saugi 19 Thailand 122 Se ISe 1 se 3 se 3 weakly sensitive
Puang Rai 2 Thailand 193 Se I Se 1 Se 3 Se 3 strongly sensitive
Sac Nau Vietnam 190 Se ISe 1 Se 3 Se 3 strongly sensitive
Progenies of Puang Rai 2/IR26760-27 were used to tag the gene Se-1(Mackill el at., 1993).

tagging genes conferring photoperiod sensitivity and critical day length are given in Table 1.
    For tagging the Se-3(t) gene with RAPD markers using the bulked segregant analysis (Michelmore et al. 1991), F3 lines derived from two F2 populations were used. These populations were: 38 F3 lines from Puang Rai 2/Nam Saugi 19 and 48 F3 lines from Sac Nau/Nam Saugi 19 studied by Poonyarit et at. (1989). Phenotyping of F3 populations of Puang Rai 2/Nam Saugi 19 and Sac Nau/Nam Saugi 19 was carried out in the greenhouse under long day length to distinguish homozygotes and heterozygotes of sensitive F3 families. The number of days to flowering of the weakly sensitive parent, Nam Saugi 19, was used as a threshold to differentiate the homozygotes and heterozygotes of sensitive groups of both crosses. Among the F3 families of sensitive group, families segregating for individuals flowering along with weakly sensitive parent (Nam Saugi 19) and strongly sensitive parents (Sac Nau or Puang Rai 2) were found. Based on the segregation of F3 families, the individual homozygous or heterozygous nature of individual F2 plant was decided. Out of 15 sensitive F3 families of the cross Puang Rai 2/Nam Saugi 19 and 23 sensitive F3 families of Sac Nau/Nam Saugi 19, a total of three and eight families, respectively, segregated for days to flowering. The F2 plants corresponding to these families were identified as heterozygous.
    Equal numbers of homozygotes of both sensitive and insensitive classes, along with the parents were used for the bulked segregant analysis with RAPDs. Of the 435 random primers (Operon Tech. Inc.) surveyed a single linked primer, A-19 was identified. This primer generated 9 bands 2 of which were putatively linked to Se-3, one associated with weak sensitivity and other with strong sensitivity. Further, the putative marker identified in Sac Nau/Nam Saugi 19 cross was monomorphic in Puang Rai 2/Nam Saugi 19 cross (Fig. 1).
    The putative markers to weak sensitivity and strong sensitivity were designated as A 19-1 and A19-S respectively. To confirm the putative linkage of the markers with phenotypes, DNA from 48 F3 DNA families were used. The marker for weak sensitivity (A 19-1) was found in 13 out of 23 strongly sensitive families of which 8 were segregating for strong sensitivity and weak sensitivity. The A19-1 was found in all the 25 weakly sensitive families. The marker for strong sensitivity (A19-S) was found in 21 out of 23 strongly sensitive families and 7 out of 25 weakly sensitive families. The linkage analysis between the respective marker and the phenotype indicated that the putative marker,



Fig. 1. RAPD markers detecting polymorphism between bulks made for alternate alleles of Se-3. PR2-Puang Rai 2, SN-Sac Nau, NS 19-Nam Saugi 19, S Bulk-strongly sensitive bulk, W Bulk-weakly sensitive bulk. Bulks were made from the homozygous progenies of Puang Rai 2/Nam Saugi 19 and Sac Nau/Nam Saugi 19 crosses. A19-I is 6.1 cM from the gene and A19-S is 12.5 cM. In both cases, recombinations between the marker and the phenotype were observed.
    The identification of a RAPD marker for the gene Se-3(t), enhancing the photoperiod sensitivity in the cross Sac Nau/Nam Saugi 19 and not in Puang Rai 2/Nam Saugi 19 indicates that the enhanced photoperiod sensitivity found in both crosses may be due to different genes and not as reported in Poonyarit et al. (1989). The phenotypic similarity observed among progenies of two different crosses cannot be taken as criterion for genetic identity since phenotypic similarity may be due to the balanced gene combination and genotype x environment interaction. From this study, it is clear that the molecular markers help to differentiate the individuals with same phenotype to have clear understanding of the genetics of complex traits such as flowering. Experiments are underway to locate the RAPD markers identified for Se-3(t) gene on rice linkage groups.
    The pre-doctoral fellowship from the Rockefeller Foundation to M. Maheswaran is duly acknowledged.

References

Kinoshita, T., 1986. Standardization of gene symbols and linkage maps in rice. In: Rice Genetics, International
        Rice Research Institute, Los Banos, Philippines pp.21.S-228.
Mackill, D. J., M. A. Salam, Z. Y. Wang and S. D. Tanksley, 1993, A major photoperiod sensitivity gene tagged
        with RFLP and isozyme markers in rice. Theor. Appl. Genet. 85: 536-540.
Poonyarit, M., D. J. Mackill and B. S. Vergara, 1989. Genetics of photoperiod sensitivity and critical day length
        in rice. Crop Sci., 29: 647-652.