Plant Breeding institute, Faculty of Agriculture, Hokkaido University, Sapporo, 060 Japan
Somaclonal variation is an important technique to generate genetic variability, for example for salt tolerance. In this experiment, the authors exploited the culture procedures to select salt tolerant callus clones. Three cultivars of japonica rice, Shiokari, Norin 8 and Taichung 65 were used. Seed-derived callus, possibly originating from the scutellum, was induced using Chu's medium supplemented with sucrose (5%) and 2.4-D (4 ppm) and subsequently transferred to the selection medium containing NaCl (0, 1.0 and 1.5%). Tolerance was evaluated by the percentage viability of the calli after 100 days from plating onto the selection medium (Experiment 1). Seed-derived calli were transferred to the B5AA suspension culture used for rice protoplast culture by Toriyama and Hinata ( 1985) (Experiment 2). The resultant calli were sieved into pieces from 0.35 mm to 1.0 mm in size and transferred to the selection medium supplemented with 0.05, 1.0, 1.5 and 2.0% NACl. After the primary selection, those calli which survived at 1.0% NaCl were shifted to NaCl-free B5AA suspension culture. The viability percentage of the resultant calli on the selection media containing 0, 1.0 and 1.5% NaCl was observed. The survival percentages of calli from Shiokari decreased prominently in higher concentrations of NaCl (Table 1).
In the primary selection of Experiment 2, calli survived up to a maximum concentraion of 1.0% NaCl. Then, suspension culture without NaCl was used for the proliferation of calli which were transferred from the medium of 1.0% NaCl. The calli of all three cultivars used in the second selection, survived even at the 1.5% NaCl (Fig. 1). When compared with the result of Experiment 1, it is evident that the primary selection increased salt tolerance of the calli. Genotypic differences among the three cultivars were also recognized. Thus, repeated selection was more effective in increasing the salt tolerance. However, it is uncertain whether the increase of tolerance is due to a genotypic change or metabolic adaptation of cultured cells. We aim to select normal, fully fertile salt tolerant lines through in vitro mutagenesis.
Table 1. Selection of calli for salt tolerance _______________________________________________________________ Cultivar NaCl in Total no. of No. survived % survival medium(%) calli _______________________________________________________________ Experiment 1: Shiokari 0 500 478 95.6 0.5 500 310 62.0 1.0 500 104 20.8 Experiment 2: Shiokari 0 250 249 99.9 1.0 250 157 62.8 1.5 250 9 3.6 Norin 8 0 250 243 97.2 1.0 250 60 24.0 1.5 250 6 2.4 Taichung 65 0 250 250 100 1.0 250 198 79.2 _______________________________________________________________
Chu, C. C., C. C. Wang, C. S. Sun, C. Hsu, K. C. Yin and C. Y. Chu, 1975. Establishment of an efficient medium for anther culture of rice through comparative experiments on the nitrogen sources. Scientia Sinica 18: 659-668.
Toriyama, K. and K., Hinata, 1985. Cell suspension and protoplast culture in rice. Plant Sci. 41:179-183.