EFFECT OF GAMMA RADIATION ON DIFFERENTIATION OF RICE CALLI UNDER ALUMINUM STRESSED

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1 EFFECT OF GAMMA RADIATION ON DIFFERENTIATION OF RICE CALLI UNDER ALUMINUM STRESSED Dameria Hutabarut* ABSTRACT EFFECT OF GAMMA RADIATION ON DIFFERENTIATION OF RICE CALLI UNDER ALUMINUM STRESSED. Calli from young spikelets of aluminum-sensitive variety Sentani were irradiated with gamma rays. After irradiation calli were differentiated on aluminum-stressed liquid medium with ph 4.\ for 4 weeks. The tolerant selection was subjected to continuous screening in R generation by using Kimura B solution enriched with aluminum, at ph 4.\ for 32 days. The alum'inum tolerance improvement was assessed as increasing in fresh weight, dry weight, plant weight, and root length. A 54 % increase of fresh weight was obtained after a dose of 20 Gy. There was no significant difference between fresh weight of plant regenerated from unirradiated callus and that from normal rice seed. ABSTRAK PENGARUII SINAR GAMMA PADA DlFERENSIASI KALUS PADI PADA MEDIA YANG MENGANDUNG AI. Kalus berasal dari mayang padi varictas yang peka aluminium, yaitu Sentani diiradiasi dengan sinar gamma. Sesudah diiradiasi kalus ditanam pad a media diferensiasi cair yang mengandung aluminium dan ph 4,1 selama 4 minggu. Seleksi ketahanan terhadap AI dilanjutkan pad a generasi berikutnya, yaitu R dcngan menggunakan larutan Kimura B yang mengandung AI, ph 4,1 selama 4 Minggu. Seleksi ketkhanan terhadap AI dilanjutkan pad a generasi berikutnya, yaitu R dengan menggunakan larutan Kimura B yang mengandung AI, ph 4,1 selama 32 hari. Kenaikan keta~ hanan terhadap AI dinilai berdasarkan penambahan berat basah, berat kering, tinggi tanaman, dan panjang akar. Penambahan sebanyak 54 % pad a berat basah didapat pada perlakuan dengan sinar gamma dosis 20 Gy. Tidak terdapat perbedaan yang nyata antara berat basah pad a R yang berasal dari kalus tanpa disinari dan yang berasal dari padi tanpa perlakuan. I INTRODUCTION In Indonesia rice is widely cultivated on lowland. Since the lowland tield has been less and less compared with the increasing need of rice as staple food, the extensification or rice cultivation seemingly to be driven to upland field. Upland soil in some areas is not yet cultivated due to its aluminum content. Aluminum is not an essential element for the growth of rice, on the contary its soluble quality to acid soil is found to be destructive. In nature, AI is chemically known as AICI3 or AI/S04)3 which is soluble in soil at a low ph. Liming is a way out to answer the soil acidity, yet it is costly. Indonesia is a tropical country with relatively high humidity so that any upland rice to be planted should be of blast tolerant variety. Sentani, the variety * Pusat Aplikasi Isotop dan Radiasi, BATAN 43

2 grown here is of that type, it gives good yield, too. But it is sensitive to AI. By applying tissue culture technique, ZAKRI (I) has been succsesfully in increasing the AI tolerance to soybean. The study was aimed to increase the AI tolerant of upland rice by a'pplying tissue culture technique. MATERIALS AND METHODS The materials used were spikelets of rice variety Sentani, about 5 to 25 mm long. The sterilized materials were inoculated on MS medium supplemented with 2 ppm 2.4-D, 1.5 g/liter casein hydrolyzate and 1.5 g/liter yeast extract. Calli were formed from swolen spikelets. After 7 days, calli were irradiated with gamma-ray at doses of 0.20 Gy, Gy, 40 Gy, and Gy and transferred to a liquid differentiation MS medium supplemented with 0.37 ppm NAA, 10.8 ppm kinetin, 1.5 g/iiter casein hydrolyzate and 1.5 g/liter yeast extract, adjusted to ph 4.1 with HCl before autoclaving. Aluminum in the form of AICI36H20 was incorporated into the medium at consentrations of 8; ; and 14 ppm. Ten ml of liquid medium was put in 500 ml bottles, so that the calli do not immerse. Calli were differentiated on Al stressed to basal medium for 4 weeks, then the plants were transffered to soil. Seeds from regenerated plants were grown to the 2 leafs stage and screened for Al tolerant using Kimura B solution enriched with ppm of AI at ph 4.1 for 32 days. The Al tolerant improvement was assessed as increasing on fresh weight, dry weight, plant height, and root length. RESULTS AND DISCUSSION Shoot Differentiation. Table 1 showed the effect of liquid differentiation medium enriched with AI, ph 4.1 upon shoots differentiation out of irradiated calli by gamma-ray. It is seen that ph 4.1 gives significant results upon differentiation of calli. In case that no AI is given, not only single formation of shoot takes place. AI given 8 ppm and ppm can stimulate the formation of shoots. While AI given at 14 ppm may inhibit the formation of shoots. Irradiated calli at dose of 20 Gy and fractionated dose Gy can accelerate the formation of shoots of calli planted on differentiation medium, containing AI or not. Even with the fractioned dose of Gy, with 14 ppm of AI can still giving a remarkable result on the differentiation of shoots: II shoots. At dose of 40 Gy and fractioned dose of Gy, was found to be a lethal dose for calli planted on differentiated medium at ph 4.1. It is seen that addition of either 8 ppm or ppm of AI was unable to stimulate the differentiation of shoots. Observation on RI' Table 2 shows the observation on R. seedling planted on solution of Kimura B enriched with ppm of AI and ph 4.1. RI tested was derivation of Ro originated from call i that treated with ppm of AI. The supporting 44

3 reason of the dose was the finding that at that treatment the unirradiated calli planted on the media having ph 4.1 can still producing shoots. This test may function as a comparison on the effect of gamma-ray upon R, seedlings. Every gamma-ray treatment on calli shows distinctive effect compared with its origin. The most remarkable effects are seen on the dry and fresh weight and the length of roots. A 54% increase of fresh weight is achieved at 20 Gy dose. There has been no significant difference on fresh weight of the unirradiated callus and its origin. Aluminum ions in a high value of ph is unsoluble so that plant can not absorb it. Maintaining its low ph is to keep its toxicity. However, with media keep at ph below 5, the agar used for solidifying the culture medium wit not gel. ZAKRI (1) used Al chelated with EDTA to maintain AI availability at a higher ph (5.8) and at the same time allow the agar to gel. In nature, there is a relationship between the toxicity of AI with low value of ph. This fact was used as a reason to chose a low ph on this study with the consequence of using liquid medium for selection. AI toxicity is indicated by inhibition of the growth of roots. AI acts directly within the peripheral root cap cells so that the mechanism of AI rice tolerante colerates with its intake absorption from the soil (2, 3). From Table 2 in seen that roots extend coincidingly with the growth of the crop itself, its longer roots its more weight. There has been no significant difference between the fresh weight of R) seedling that of calli and that of its origin (Sentani) is seeds, however the different length of their roots are highly significantt (Table 2). CHOUDRY et al. (4) and OKAWARA et at. (5) state that the growth of roots may serve as good indicator to the AI tolerante of cultivar. Table 2 shows that irradiated callus is an effective treatment to increase the AI tolerance of rice. CONCLUSION 1. A combination of tissue culture technique and gamma-ray treatment is applicable to increase the Al tolerant of rice. 2. Irradiated callus with gamma-ray at either 20 Gy or fractionated Gy dose can accelerate the shoots differentiation out of callus planted on a liquid differentiation medium with ph 4.1, containing AI or not. 3. A 54% increase of fresh weight is acheived by R. seedling that irradiated by 20 Gy of gamma-ray and Al stressed as much as II ppm. ACKNOWLEDGEMENTS Author wish to thank Mr. Parno for his kind help. 45

4 REFERENCES t. ZAKRI, A.H., "Induction and selection of aluminum resistant variants from soybean celt culture for plant improvement", IAEA Symposium (1986) BENNET, RJ., BREEN, C.M., and FEY, M.V., The effects of aluminum on root cap function and root development in Zea mays L. Env. Exp. Bot. 27 (1987) FEGERIA, Influence of AI in nutrient solution on chemical composition in 2 rice cultiyars at different growth stages. Rice Abstr. 2 6 (t 986) CHAUDRY, M.A., YOSHIDA, S., and VERGARA, B.S., Induced mutations aluminum tolerance after N-Methyl-NN-Nitroso urea treatment of fertilized egg celt in rice. Env. Exp. Bot. 27 (1987) OKAWARA, R., OJIMA, K., and YOSHIDA, S., Selection and planted regeneration (1987) t aluminum tolerant caltus derived from rice grain. Rice Abstr. 1Q 4 46

5 Al, Gamma-ray Table The liquid of effects 1. callino. of upon ph inoculated differentiation (mgjl) Total Al 4.1, shoot 14 regenerating dose of 1 No. shoots differentiation medium of enriched out shoots 0 Table 2. The observation on R1 seedlings planted of Kimura B enriched with AlII ppm and ph 4.1 (Each figure repre sents the mean of 210 seedlings) provious dose of gamma-ray (Gy) Provious dose of Al (ppm) Fresh weight (mg) Dry weight (mg) Shoot lenght (mm) Root length (mm) o Origin (Sentani) c. v. (%) L.S.D. at 5% level 1% level

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