Mutation Breeding of in Vitro Research Proposal

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The size of the petiole can be increased or decreased through use of gamma radiation.

XI. The Effect of Mutation Breeding on Shoots and Stem

The initiation of shoots is controlled by the large number of genes in higher plants. This is clearly demonstrated in the alternations in shoots resulting following treatment with a mutagen. The increase in the number of shoots has many benefits which includes the increase in the number of branches which can ultimately result in the increase of yield according to Savov (1983). Furthermore, the proportion of shoots to the length of internodes is an essential factor of plants and in Malus pumila treated with gamma radiation demonstrated is an improvement in shoots and internodes ration. Therefore, the growth of the plant resulted through an increase in the number of branches. (Paprstein, 1988)

XII. The Effect of Mutation Breeding on Plant Height

A change in the height of a plant is an important horticultural characteristic however, plant height in mutation breeding is not a common feature as tallness in a plant is not a desired characteristic since an increase in the height of a plant results in a negative impact on the plant stem stability.

XIII. Additional Morphological Information

The work of Arora, Nakao and Nakajima (1970) entitled "Perpetuation of Begonia Rex by Aseptic Culture with Micro-Leaf Cuttings Under Various Conditions of Auxin and Cytokinin" state that they conducted a study in which micro-leaf cutting of Begonia rex were aseptically cultured with the aim to isolate and propagate the irradiation induced chimeras." (p.275) Micro-leaf cutting of Begonia rex are stated to have regenerated roots and a few buds when they were cultured ascetically on White's basal medium. It is stated that the lower concentrations of NAA "...showed stimulatory effect on root initiation which was observed 35 days following inoculation. Callus tissue was formed by the varying concentration of NAA microcultures and the maximum number of roots per rooted culture is stated to have been "recorded at 0.1ppm of NAA." (Arora, Nakao and Nakajima, 1970, p.276)

It is however reported that the lowest concentration of NAA as well as in control where callus did not form at all, in a few cultures a few buds were formed whereas as relatively higher concentrations bud formation was completely suppressed. No root formation took place during the thirty day period when the temperature maintained was between 17~22 degrees Celsius however following one month of the temperature being increased to 27~30 degrees Celsius most of the culture began regeneration of roots.

It is reported that the combination with the 0.1 ppm NAA that the maximum percentage of culture regenerated roots on the 25th day however in the combination with 9.91ppm NAA organ differentiation was constant until the 35th day however there was a sudden increased noted up until the 50th day.

Arora, Nakao and Nakajima state that a "number of species in the genus Begonia are well-known for their ability to form both adventitious buds and roots on detached leaves without the aid of plant regulators." (1970, p.280) It is reported that Heide (1965) reported that the "stimulatory effects of these substances in the asceptic culture of Begonia with expanding leaves of about 3.5 cm diameter along with petioles trimmed to a length of 2cm and it is stated that the results of the experiment reported in the work of Arora, Nakao and Nakajima provide explanation that in the asceptic propagation of Begonia with micro-leaf cuttings of 3X3 mm auxin and cytokine are needed to produce speedy and better generation of organs." (Arora, Nakao and Nakajima, 1970, p.280)

In fact, it has been reported in previous studies including those of Heide (1965) and Wirth (1960) that B. rex requires a high cytokinin to auxin ratio for bud formation to be stimulated and root formation to be suppressed. A low ratio of cytokinin to auxin ratio has the opposite effect in that root suppression does not occur and bud formation is not stimulated. NAA at a high concentration further promoted the development of roots and inhibited the formation of buds while the high concentration of kinetic in turn promoted formation of bud to the maximum degree and almost but not quite inhibited root formation. It is however reported that the same combination with auxin and cytokinin in equal concentration and the difference in the percentage of roots and buds of cultures can be credited to the developmental stage of the mother plants and the leaves' age due to the fact that during collection of leaves it was not possible to precisely determine the ages of the leaves.

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NAA being applied following kinetic resulted in the production of better differentiation of organs than the two combined. Buds were regenerated by cultures on kinetin media alone at a low range of temperature (17~22 degrees Celsius) however the majority of cultures took one month for regeneration of buds and only when the temperature range was increased to 27~30 degrees Celsius with culture also regenerating roots.

It was further noted that Heide (1965) reported "in Begonia that high temperature suppresses bud formation and counteracts the promotive effect of cytokinins on this process and the inhibitory effect of cytokinin on root formation." (Arora, Nakao and Nakajima, 1970, p.281) Therefore it is stated to be evident that "for early differentiation of organs with better growth in the multiplication of B. rex by aseptic culture with micro-leaf cuttings, auxin and cytokinin are required in a defined ration or separately at certain stage of organ development under favorable temperature conditions." (p. 282)

High success is possible to achieve when "micro cuttings are first cultured on kinetin medium and later transferred to NAA medium where almost all of the cultures regenerated roots as well as buds with better growth." (Arora, Nakao and Nakajima, 1970, p.282) The most effective for speedy regeneration of organs is stated to be a combination of NAA (0.01ppm) and kinetin (0.1ppm)." (Arora, Nakao and Nakajima, 1970, p.283) In addition, this method may be applied with the benefits of isolation and diversion of irradiation induced chimeric tissues into fully homogenous plants." (Arora, Nakao and Nakajima, 1970, p.284)

The article "Genetic Manipulation In Crops: Proceedings of the International Symposium on Genetic Manipulation in Crops, the 3rd International Symposium on Haploidy, the 1st International Symposium on Somatic Cell Genetics in Crops, Beijing, October 1984" reports that the use of colchicine in culturing the explants of Begonia rex were successful when propagated in vitro. Furthermore, "the plantlets of Begonia rex...with fancy leaves cultures in vitro remained true to their original types." (1984) It is reported that Lindemuth (1904) "removed and rooted mature begonia leaves and observed that they then increased considerably in size, chiefly because of cell enlargement. While they were attached to the plant this presumably was prevented by "correlative inhibition." Similar results have been reported by others." (3rd Symposium on Haploidy, 1984)

It is reported that in the leaves of Begonia "the portions on either side of the midrib are usually quite unlike; and there are marked internal asymmetries, as when two daughter cells are dissimilar. In most cases of this sort, however, the asymmetry proves to be part of a larger and more complex pattern which is symmetrical. Symmetry is often more conspicuous in embryonic structures or meristematic regions than at maturity, and some of its most remarkable examples. Neel (1940) reports that in Begonia where there are "...silver spots on the leaf surface, there is a hair formed in each spot save the very small ones, and the larger the spot, the longer the hair." (cited in: 3rd Symposium on Haploidy, 1984)

It is reported that root and shoot restoration on leaves which have been removed from the plant and in which petiole has been placed in the soil has occurred and that in such cases "...after adventive roots and buds have been formed on lamina or petiole, various anatomical changes may be observed, especially a marked increase in the vascular tissue of the petiole. Furthermore, instead of being disposed in an arc, as in normal petioles, this tissue often enlarges to form a complete vascular ring. The petiole thus becomes structurally as well as functionally a stem. Simon (1929) is stated to have noted this in the Begonia and stated that the "...bundles from the young roots induced new vascular tissue only in that part of the petiole just below them, suggesting the polar distribution of a hormone. Similar results were obtained by Doyle (1915) through grafting buds onto the petioles of rooted leaves." (3rd Symposium on Haploidy, 1984)

The location of the regenerated structures is determined mainly by the anatomy of the leaf." However, it is important to note that in some species, including that of Begonia rex that while not always succulents "...restoration and vegetative reproduction occur in other parts of the leaf than the base." (3rd Symposium on Haploidy, 1984) In Begonia rex it is stated that "...shoots may be induced very readily on petiole and….....

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