Study on breeding of Cucurbitaceae using biotechnology

The successful application of biotechnology such as cell fusion and transformation of the breeding of Cucurbitaceae crops, which are known to be recalcitrant in tissue culture, was made possible from the use of improved tissue and cell culture techniques. The present study dealt with investigations on the various factors and conditions involved in plant tissue and cell culture techniques of Cucurbitaceae. Firstly, conditions that influence regeneration from plant tissue was investigated in melon, cucumber and watermelon. It was proven that cotyledonary explants of mature seeds and young seedling possess high ability for shoot organogenesis and embryogenesis. Moreover, shoot organogenesis and embryogenesis of melon and cucumber were found to be controlled by concentrations of auxin in regeneration media. Secondly, regeneration system from protoplasts of melon was also developed and then applied to the cell fusant obtained between melon and wild relative. To analyze hybridity, a novel analysis method, called DIG-RAPD, was developed. The method was more sensitive than the other conventional methods. Somatic hybrid calli produced by symmetric cell fusion between melon and cucumber exhibited spontaneous asymmetric hybrid nature suggesting that many genes of one parent were eliminated during cell division. Thirdly, transformation system in cucumber was developed and then applied to the integration of rice chitinase gene, a disease resistance gene,into cucumber. The transgenic cucumber exhibited increased resistance against gray mold. Finally, the influence of transgenic melon introduced with coat protein gene of cucumber mosaic virus on environment was evaluated in a close and a semi-closed greenhouse. The results of this study provided solutions to the major problems encountered in the application of cell fusion and transformation to breeding of cucurbitaceae