41. Callus induction and growth from different Oryza species


Koh-ichi MORI

Plant Breeding Institute, Faculty of Agriculture, Hokkaido University, Sapporo, 060 Japan


The problems involved in the successful use of the protoplast culture method in rice breeding, namely, stable isolation of protoplasts, protoplast culture, and selection of fused cells, remain unsolved. The anther culture of rice was pioneered by Niizeki and Oono (1968) who obtained haploid and diploid plants from microspores. This technique was used to obtain promising true-breeding lines from F\1\ plants at Hokuriku National Agricultural Experiment Station and Kamikawa Agricultural Experiment Station of Hokkaido, in Japan. Niizeki nand Kita (1981) succeeded in obtataining fusion of rice and soybean protoplasts by using polyethylene glycol (PEG)-high pH- high Ca++ techniques, but the culture of fused protoplasts and regeneration of plants were unsuccessful.

I attempted to induce callus formation from seeds of 40 strains belonging to 9 Oryza species and studied the proliferation of the calli. The seeds were surface-sterilized by immersion in 10% bleach and were rinsed three times in sterile water. Then, Chu's medium containing 2 x 10-5 M 2,4-D was used for seed callus initiation. The callus growth was maintained by subculturing repeated every 10 days. Callus induction and proliferation were observed 40 days after plating.

All species except O. latifolia produced seed calli, but they clearly differed in callus induction and proliferation ability (Table 1). The seed calli from O. sativa and O. glaberrima showed better proliferation than those from other species. In O. sativa, subspecies japonica showed better proliferation than indica. A wide range of variation in callus growth or proliferation was also observed among japonica strains, suggesting that callus proliferation is under genetic control. The wild species generally showed a poor callus growth, the diameter of calli being nearly 0 to 3 mm after 40 days of incubation.

The techniques for protoplast fusion and culture of hybrid protoplasts are being studied. It seems necessary to screen the genotypes for a highly prolific callus growth and to obtain suitable media containing optimum levels of growth regulating substances for successful cell breeding.


Table 1. Comparison of callus growth among Oryza species

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                                        Diameter of callus (mm)

       Species name    No. of                                        Mean
                       strains  0  1  2  3  4  5  6  7  8  9  10
=============================================================================
sativa                    18                1  3  6  1  3  2   2    6.9
   subsp. japonica        14                   2  4  1  3  2   2    6.6
   subsp. indica           4                1  1  2                 5.1
glaberrima                 5                1  1  2  1              5.7
punctata                   4       1  2  1                          1.9
minuta                     1       1                                1.2
officinatis                6       3  3                             1.2
australiensis              2             2                          3.2
latifolia                  1    1                                   0.0
grandiglumis               1             1                          2.8
brachyantha                2    2                                   0.1
=============================================================================
Note: Callus diameter after 40 days from plating

References

Niizeki, Hiroo and K. Oono, 1968. Induction of haploid rice plants from anther culture. Proc. Japan Academy 44: 554-557.

Niizeki, Minoru and F. Kita, 1981. Cell division of rice and soybean and their fused protoplasts. Jpn. J. Breed. 31: 161-167.