Study of Breeding of Mulberry by Tissue Culture

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New breeding of mulberry used a tissue culture laying stress on a protoplast fusion was tested by some basic experiments. At first, we investigated a preservation method of callus sample. Then, it was confirmed that a cold storage method (st 5^ﾟC) after pretreatment made the preservation possible up to 20 weeks. In this cold storage test, we found of exist a similar phenomenon as dormancy in the cold staged cellus. It became clear that this phenomenon began after 3 weeks and became the most marked after 8 weeks, but after 18 weeks they awaked from this phenomena and moved to the state just as dormancy, caused by the condition of the outer field temperature. It appeared that all dormant mulberry callus had this character but the mulberry with difficult dormancy (for example Okinawa mulberry etc.) had not. Enzyme and protein of callus at shipping were analyzed by the electrophoresic method. The rise and fall of their bands in period of the cold stage had something in common with that of the similar phenomena as dormancy, which broken down by adding gibberellin and dithiothreitol to the culture medium. With a freezing preservation for long time storage, further, we develod more simple freezing preservation method by a refregiration than an ordinary liquid nitrogen method by our investigation of pretreatment drug and method. Next, the collosion isolation method of sample cell was studied by using the growth regulatory substances. By protoplast electric fusion of the abovementioned cell, it is recognized to exist the difficult fusion between mulberry varieties. With attempting the fusions between them and the foreign plant, they were almost succeeded, but some of them were broken by high voltage. We improved the ordinary composition of culture medium and atta ined tothe inducible growth from adventitious embryo to adventitious root. We are rearing them now. From these results, we can report to attain our purpose of the original plan.