25. Genetic analysis of some traits of paddy and upland rices using doubled haploids derived from the F1 plants of Koshihikari X Sensho

Ikuo ANDO 1, Taiji YOSHIDA1 and Naoki KISHIMOT02

1) Department of Crop Development, National Agricultural Research Center, 2-1-2 Kannondai, Tsukuba, Ibaraki, 305 Japan.

2) Department of Molecular Biology, National Institute of Aerobiological Resources, 2-1-2 Kannondai, Tsukuba, Ibaraki, 305 Japan.

It is considered that most of agronomic traits are controlled by quantitative trait loci (QTL). It is also known that doubled haploids (DH) are useful tools for analysis of QTL (Thompson et al. 1991). By using DH lines derived from the F1 plants of Koshihikari (paddy rice) X Sensho (upland rice), we attempted to map the following traits: the horizontal resistance to rice blast and the resistance to rice stripe derived from Sensho, and good eating quality from Koshihikari (Table 1).

Table 1. Comparison of three traits between Koshihikari and Sensho

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Trait                            Koshihikari         Sensho
===================================================================
Field resistance for blast       low level           high level
Resistance for rice stripe       susceptible         resistant
Eating quality                   good                poor
===================================================================

Development of DH lines : After the anthers from two F1 plants were cultured, 101 diploids were obtained from regenerated plants.

Test for the horizontal resistance to rice blast : The DH lines were sown in the upland field where the race 037.3 of blast, Magnaporthe grisea, was introduced artificially. The disease severity of the DH lines was scored using 11 grades from 0 (no lesion) to 10 (all plants are dead).

Test for the resistance to rice stripe : In a net cage, the DH lines (2.5 leaf age) were exposed to viruliferous small brown plant hoppers, Laodelphax strialellus. The resistance of the DH lines was evaluated according to Sakurai et al. (1963).

Test for eating quality of boiled rice : Eating quality was evaluated by measuring the glossiness of boiled rice (Fujimaki and Kushibuchi 1975) that was scored using 6 grades from 1 (bad) to 6 (good).

Restriction fragment length polymorphism (RFLP) analysis : DNA studies were conducted according to Saito et al. (1991). The total DNAs isolated from mature leaves of each parent and 70 DH lines were digested with four restriction enzymes, BamHI, Bgl II, EcoRV and Hind III. A total of 16 RFLP markers were selected from chromosome 4 and 11 in which two genetic markers, Ph and la, are located respectively, based on their positions on the RFLP map previously published (Saito et al. 1991; Kishimoto et al. 1993). It is known that Ph and la are linked with the horizontal resistance to blast in previous studies (Ph: by Maruyama et al. 1983; la: by Higashi and Saito 1985).

The segregation for the horizontal resistance to rice blast (Fig. 1) and the glossiness of boiled rice (Fig. 2) showed continuous distribution that has a peak between the parents. As for the segregation of resistance to rice stripe disease, the ratio of resistant DH lines to susceptible lines was about 1:3 (Table 2). This ratio shows that the resistance to rice stripe derived from Sensho is controlled by two loci. There was no significant correlation among these three traits by t-test. Of the 16 RFLP markers selected, 6 markers showed polymorphism between the parents. Four polymorphic markers were used for the RFLP analysis of DH lines (Fig. 3). The segregation ratio of these 4 RFLP markers and Ph was 1:1. In the DH lines that showed the genotype of Sensho at the locus of Ph or Xnpb267 (on chromosome 4), the disease severity for blast was significantly lower than that in the DH lines which showed the genotype of Koshihikari (Table 3). In the DH lines that showed the genotype of Sensho at the locus of XNp181 or XNpb320 or XNpb2O2 (on chromosome 11), the ratio of resistant lines to rice stripe was significantly higher than that in the DH lines which showed the genotype of Koshihikari (Table 3). The results summarized in Table 2 indicate that the resistant genes to these two diseases exist near these 5 markers mentioned above (Fig. 4). There was no significant correlation among these 5 markers and the glossiness of boiled rice.

Table 2. Reaction of DH lines to rice stripe

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R     M*     S      Total     Expected ratio     X2        P
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19    7     72      98 (91)         1:3          0.93  0.30-0.50
================================================================
  * M type lines which showed the intermediate reaction were
excluded in the X2 test.

Table 3. Correlation between three traits and five genetic markers

========================================================================
Markers      Genetic type        Bl.        St.                Gl.
                                            R:S
========================================================================
Ph            K-type             5.0 **    12:37  ns.         3.1  ns.
                                    |            |                |
              S-type             3.1        7:33              2.9
========================================================================
XNpb          K-type             4.8 **     8:28  ns.         3.0  ns.
                                    |            |                |
267           S-type             3.0        6:16              3.0
========================================================================
XNpb          K-type             4.4 ns.     4:26 **          3.0  ns.
                                    |            |                |
181           S-type             4.0        10:20             3.0
========================================================================
XNpb          K-type             4.0 ns.     1:37 **          3.0  ns.
                                    |            |                |
320           S-type             3.8        13: 9             3.0
========================================================================
XNp           K-type             4.2 ns.     0:37 **          3.0  ns.
                                    |            |                |
202           S-type             4.0        14: 8             2.9
========================================================================
               Bl.-Blast, severity grade: 0-10 (all dead).
               St.-Rice stripe, reaction grade.
               Gl.-Glossiness level of boild rice: 1 (bad)-6 (good).
               K-type: Koshihikari type, S-type: Sensho type.
               Significant at 1% level, ns.-not significant.




References

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Kishimoto, N., M.R. Foolad, E. Shimosaka, S. Matsuura and A. Saito, 1993. Alignment of molecular and classical linkage map of rice, Oryza sativa L. Plant Cell Reports 12: 457-461.

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Sakurai, Y., A. Ezuka and H. Okamoto, 1963. The seedling test method of varietal resistance of rice plants to stripe virus desease (Part 1). Bull. Chugoku Agr. Exp. Stat. A 9: 113-125. (in Japanese with English summary)

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