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PROBLEMS IN USING INDUCED MUTATIONS IN VEGETATIVELY PROPAGATED PLANTS K.Mikaelsen* ABSTRAK - ABSTRACT PROBLEMS IN USING INDUCED MUTATIONS IN VEGETATIVELY PROPAGATED PLANTS. Conventional breeding techniques are difficult to apply in most vegetativdy propaga- ted plant species. The mutation breeding is therefore very important in these plant species, but practical results have been difficult to obtain due to the problems in recovering the mutated cells in the plant. The importance of various techniques are discussed and the advantage develop- ing various tissue culture techniques have been described. MASALAH DALAM PENGGUNAAN MUTASI BUATAN PADA TANAMAN YANG DIBIAK SECARA VEGETATIF. Teknik pemuliaan secara konvensional umumnya sukar diterapkan pada spesies tanaman yang dibiaksecara vegetatif. Oleh karena itu teknik pemuliaan mutasi sangat penting bagi spesies tanaman yang demikian, tetapi mutan yang dapat digunakan dalam praktek sukar diperoleh karena masalah kembalinya sd yang termutasi dalam tanaman ke bentuk normalnya. Dalam makalah ini dibahas keunggulan berbagai teknik mutasi buatan clankeuntungan pengembangan berbagai teknik kultur jaringan. INTRODUCTION A large number of plant species are propagated asexually (or vegetatively) such as many root and tuber crops, sugarcane, fruits, and other three species and shrubs and many ornamentals. Cross-breeding is often limited by specific problems in most vegetatively propagated plants (VPP). Many VPP have a rather long vegetative phase before going into sexual reproduction. In addition, these plants are generally highly heterozygous, which causes complicated segregations and makes the detection of a useful recom- binant very difficult. 'This problem is further enhanced by frequently polyploidi in such plants. All these factors make cross breeding very difficult and time consuming. Furthermore, incompatibility and other cross barriers, apomixis and sterility exist quite often and hinder the plant breeder in making use of conventional cross-breeding. * Expert, UNDP Project INS/78/074 107 MUTATION BREEDING OF VEGETATIVELY PROPAGATED PLANTS' It is, therefore, quite clear that mutation breeding techniques are an attractive alternative in breeding of VPP. The most promising aspect of mutation induction in VPP compared to cross-breeding methods is the ability to change only a few characters of an otherwise good cultivar without altering significantly the original well established genotype. Mutation breeding, therefore, must be considered as the obvious means to perfect the leading cultivars and as a possible sh~r_tcut for inducing desired genetic alterations in outstanding cultivars. Obviously, mutations are the only means for producing variability in sterile VPP and in obligate apomicts. Further, mutations can be useful to break apomixis, to overcome self-sterility and cross barriers as well as to uncover rearrange chimeras. In spite of these many advantages of mutation breeding techniques in VPP the results have been rather small in comparing with seed propagated plants such as rice, barley. and many others crops. The main problem has been the recovery of mutants in VPP. Normally a shoot, or any other multicellular organ are treated for mutation induction and the desired mutations has occured in one cell. The chances of such a mutated cell growing into a sector or layer and be able to manifest itself will depend on l!s .position within the apex as well as its growth rate as compared with the surrounding cells or tissue layers. It is obvious that mutated cells will be easily hidden and not be visible in such multicelledar tissues and special methods are required to unciver such mutated cells. The first requirement is, therefore, that mutagen treated (unirradiated) material has to be propagated to permit the formation of periclinallayers before selection can be applied. Very often repeated propagations are necessary for obtaining large sectors of mutated tissue from which a maximum number of mutants can be recovered. In fruit trees, for example, (Figure 1) the best conditions are found in the axillary meristems of the basal leaf primordia of the dormant buds. (The primordia to be used for treatment should consist of as few cells as possible). Experience shows that the buds derived from leaves 4 - 8 (approximately) of the primary shoot are those that exhibit the highest frequency of relatively broad mutated sectors in the second vegetative generation. Measures have to be taken (by pruning) to force these buds into growth. Several authors have reported better results after irradiating buds that have just started growth than with buds in deep dormancy. If growing plants are irradiated, decapitation of the main shoots will force new buds to develop through regeneration and thereby increase the chance for recovering a larger number of mutants. Many of the complications described above could be eliminated if chimera formation could be avoided. One of the methods available is the adventitious bud techniques which was developped by BROERTJES and collaborators at Wageningen (1). This technique is based on the phenomenon that the apex of the adventitions buds, such as may be formed the base of the petiole of detached leaves, originates from only one (epidermal) cell. Consequently, adventitious plantlets either are completely normal or are complete, solide mutants. In other words, chimera formation does not take place. This adventitious bud technique offers a great 108 advantage for practical mutation breeding and has given many good results particu' larly in ornamental plant (Saintpaulia, Streptocarpus, Achimenes, Begonia, Liliwrli and many others). Many plant species can be propagated this way. BROERTJES ~~. (1) list other 350 of such species. Many species of economically important plant families have not been tested for developing the adventitious bud techniques and may not always develop adventitious buds from only one cell. In addition, the condition of the mother plant, the age of leaves, environmental conditions during and after rooting and the auxin-cytokinin balance appear to influence the formation and differentiation of adventitious buds. POTENTIALS OF TISSUE CULTURE TECHNIQUES IN BREEDING OF VPP The modern developments of culturing somatic plant cells should also be investigated for use in VPP in this respect tissue culture technology holds much promise. Recent advances in the field of plant protoplast, cell, tissue, and organ culture (embryo, anther) have, transferred this area if fundamental research into one that is dynamic and promising also for obtaining further advances in crop improvements programmes. I believe the potentials of this technology can be powerfull tool also in the VPP. During the last 30 years, the culture of ovule, ovary, and embryo has been employed to overcome sterility. incomtability and dormancy; to induce poly· embryony; and to succesfully hybridize various crops. By meristem culture, large numbers of horticulturally important plants have been commercially propagated and freed from pathogens. These established techniques have played an important role in wide hybridi- zation and clonal propagation programmes and will, no doubt, continue to contri- bute to future demands. However, some of the recent advances in the area of plant protoplast, cell, tissue, and organ culture have attracted international attention because of their significance in and far-reaching implications for agricultural research and crop improvement programmes. It is expected that the following techniques will playa significant role, not only in the improvement of existing crops but also in synthesizing new plants : 1. Wide hybridization in crops through in vitro pollination and fertilization. 2. Production of haploid and homozygous plant from exised anthers, isolated pollen and by chromosome elimination. 3. Somatic hybridization and genetic engineering through the fusion of protoplasts and the uptake of DNA (Figure 2). 4. Induction of genetic variability in crops (mutants, and various ploidy levels) through protoplast and cell culture and selection in these cultures for resistance to diseases, salinity, water, and temperature stresses and possibly many other important characteristics. I expext that the VPP will have great potentials for benefits from such techniques. 109 REFERENCES 1. BROERTJES, C:; HACCIUS, B., and WEIDLICH, S., Adventitions bud for: mation on isolated leaves and its sigriificance for mutation breeding, Euphytica 17 (1968) 321. 2. DONINI, B., '1nduction and isolation of somatic mutations in vegetatively propagated plants", Improvement of Vegetatively Propagated Plant Through Induced Mutations (Technical Reports Series No. 173), IAEA, Vienna (1975)35. 3. BAlAl, Y.P.s., Potential of protoplast culture works in agriculture, Euphytica 23 (1974)633. 4. CARLSON, P.s., SMITH, H.H.,and FEARING, R.D., "Parasexual interspecific plat hybridization", Manual on Mutation Breeding, 2nd Ed. (Technical Reports Series No. 119), IAEA, Vienna (1977) 206. 110
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