| Muskmelon (Cucumis melo L.) and watermelon (Citrullus L.) belong to Cucurbitaceae. They are the fruits of high economic value, widely distributed in the world and have a wide range of cultivars. However, during the growth of muskmelon and watermelon, they were often affected by pest and disease, especially fusarium wilt and anthracnose, which seriously impact on the yield and the quality of muskmelon and watermelon. Many fine cultivars have been bred by conventional breeding methods, but the improvement of certain traits is difficult. Using genetic engineering to improve muskmelon and watermelon has become a direct and effective means. However, high efficient tissue culture and plant regeneration technology is a prerequisite for transgenic technology.Antimicrobial peptide protein which was extracted from Ginkgo biloba has strong resistance to some melon fungi like Fusarium and anthracnose. In this research, the high efficient genetic transformation system of muskmelon and watermelon was studied, the fast and high effectient regeneration and genetic transformation system was established, then antibacterial peptide gene Gnk2-1 was transferred into melon and watermelon by agrobacterium-mediated transformation, to breed new resistant varieties by molecular breeding. The main results were as follows:1. The high efficient regeneration system of muskmelon was established. The cotyledon explants of muskmelon FY1,HY2 were used as the experimental material to study the influence of different hormone combination on the regeneration system of the melon. The results of the study showed that the reproducibility of the cotyledon explants of FY1 is higher than that of HY2 under the same condition, the optimal medium for the FY1 cotyledon callus and adventitious bud induction was MS+6-BA2.0mg·L-1+IAA0.1mg·L-1, and the inductivity was above 95%; the most suitable medium for the buds to lengthen was MS+KT0.2mg·L-1; the best medium for root induction was 1/2MS+IAA0.1mg·L-1.2. The high efficient regeneration system of watermelon was established. The cotyledon explants of watermelon 134-2 Biao,Jing-fu-2 were used as the experimental material to study the influence of different hormone combination on the regeneration system of the watermelon. The results of the high efficient regeneration system in watermelon cotyledon showed that the optimal medium for the 134-2 Biao cotyledon callus and adventitious bud induction was MS+6-BA2.0mg·L-1+IAA0.2mg·L-1, and their inductivity were both above 96% which was higher than that of Jing-fu-2; the most suitable medium for the buds to lengthen was MS+KT0.2mg·L-1; the best medium for root induction was 1/2MS+IAA0.1mg·L-1.3. The melon and watermelon genetic transformation system was optimized. The cotyledon of muskmelon FY1 and watermelon 134-2 Biao were used as the explants. The concentration of Kanamycin was selected, Kanamycin sensitivity test proved that Kanamycin had an apparent inhibition to the growth and the differentiation of melon and watermelon explants, the appropriate selection pressure was 100mg·L-1. The cotyledons were precultured for 2d, when the bacterial concentration is about 0.5 OD600, the explants were infected for around 10min, then the explants were cultured with the bacterial for 3d, and the conversion efficiency was the highest. The influence of Acetosyringone (AS) on the conversion efficiency of melon and watermelon was not manifest.4. Antibacterial peptide gene Gnk2-1 was transferred into melon and watermelon by agrobacterium-mediated transformation. One positive transgentic plant of muskmelon was obtained by identification of PCR. The detached leaves of transgenic plants were inoculated with the Fusarium, and the plant had a stronger resistance to Fusarium wilt. |
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