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    Several reports are available on induction of variation in agronomically important characters through tissue culture techniques in rice (Onno 1981; Ogura et al. 1987; Abdullah et al. 1989; Li and Murai 1990). These findings have aroused considerable interest with regard to possible implications of in vitro induced variations for plant breeding and interpretation of nature of the variations. In this paper we report a study on agronomically important variations in the regenerants from protoplast (protoclone) and microspores (pollenclone) of cold tolerant rice cv. RCPLI-IC.
    Microspore derived calli were obtained from cold tolerant rice cv. RCPL1-1C on E24 semisolid medium as described by Bhuyan et al. (1990). These calli were maintained till ten months by regular subculturing on N6 medium supplemented with 4.0 mgl -1 2,4-D, 20 gl -1 sucrose, 30 gl -1 mannitol and 8 gl -1 agar. The calli derived from protoplasts were obtained from eight months old microspore derived cell suspensions of cold tolerant rice cv. RCPL1-1C as described by Gupta etal. (1993).
    The calli obtained from both sources, i.e., microspores and protoplasts were transferred onto MSB medium (MS salts with 3.0 mgl -1 kinetin, 0.5 mgl -1 NAA, 30 gl -1 sucrose and 8% agarose) for differentiation and plant regeneration. The cultures were kept initially in dark for one week to facilitate diffferentiation and then transferred to a 16/8 h light/dark regime using an assortment of fluorescent light (3000 lux) at 26 ± 1°C. At two to three leaf stage, the developing plantlets were transferred to MS/2 semisolid medium for root development. After the establishment of root system, the plantlets were transferred to MS/2 liquid medium without sucrose for hardening and then transferred to pots containing 1:1 sterilized soil and compost. The seeds of RCPLI-IC were grown in pot and treated as a control in this experiment.
    The data related to yield contributing character viz., plant height (cm), flag leaf length (cm), flag leaf width (cm), flag leaf length/width ratio, (flag leaf of main tiller), number of productive tiller, panicle length (cm), number of spikelets per panicle and 1000 seed weight (g) were recorded at maturity of plants. The means of protoclones and pollenclones were compared by 'paired t test'.
    The regenerants of pollenclones exhibited wide range of variation in plant height, flag leaf length, flag leaf width, flag leaf length/width, number of productive tiller, panicle length and number of spikelets per panicle compared to protoclones (Table 1). Perhaps, the variation within pollenclones, was more due to lack of equal exposure of each regenerating cells with medium during subculture whereas in protoclones, each regenerating cells were equally exposed with medium.
    The significant variation was observed in plant height, flag leaf length, flag leaf width, flag leaf length/width, number of productive tillers, number of spikelets per panicle and 1000 seed weight in protoclones as compared to control. The pollenclone exhibited

Table 1. Mean and standard error of different characters in protoclones and
            pollenclones of cold tolerant rice cv. RCPL 1-1C (N=15)
 

Table 2. Comparision of means of characters studied by paired 't' test
 

At 5% level of significance.
S: Significant.
NS: Nonsignificant.
 

significant variation for flag leaf width, flag leaf length/width, panicle length and 1000 seed weight compared to control. However, both sources produced bold seeds compared to its control (Table 2). The comparision between pollenclones and protoclones, the protoclones exhibited significant variation, for more number of characters and the variation for plant height, flag leaf length, flag leaf width, flag leaf length/width, number of productive tiller, panicle length and 1000 seed weight was significantly different to pollenclones (Table 2). Perhaps, the variation in protoclones was more due to their exposure to several media as well as cultural conditions, i.e., AA medium, enzyme mixture, heat shock treatment, CPW solution and modified N6 medium, compared to pollenclones which have undergone culture only through modified N6 medium.

References

Oono, K., 1981. In vitro method applied to rice. In Thorpe, T. A. (ed.) Plant tissue culture. Academic Press,
        New York, pp.273-298.
Ogura, H., J. Kyozuka, Y. Hayashi, T. Koba and K. Shimamoto, 1987. Field performance and cytology of
        protoplast derived rice (Oryza sativa L.): High yield and low degree of variation of four Japonica
        cultivars. Theor. Appl. Genet. 74: 670-676.
Li, Z. J. and N. Murai, 1990. Efficient plant regeneration from rice protoplast in general medium. Plant Cell.
        Reports 9:216-220.
Bhuyan R. N., J. N. Gupta, A. Paltanayak and H. S. Gupta, 1990. Callus induction and plant regeneration from
        microspores of indica rice. Proc. International Symposium, Rice Research: New Frontiers. 15-18 Nov.
        1990, Hyderabad, India, pp. 127-129.
Gupta J. N., A. Pttanayak and H. S. Gupta, 1993. Plant regeneration from protoplasts of cold tolerant rice
        varieties. RGN. 10:113-116.
Abdullah R., J. A. Thompson, G. S. Khush, R. P. Kaushik and E. C. Cocking, 1989. Protoclonal variation in the
        seed progeny of plnats regenerated from rice protoplasts. Plant Science 65: 97-101.
 
 

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