The Research Of Agrobacterium-mediated Genetic Transformation System Of Watermelon

China is the leading producer and consumer of watermelons in the world. Butconventional breeding process is long and laborious, so germplasm innovation andsupply of new watermelon varieties can not meet market demands. Geneticengineering has become an effective tool of breeding new varieties of watermelon toimprove its quality．With the nutrition and health function of carotenoids beingwidely recognized in recent years, more and more scholars investigate watermeloncarotenoid composition and biosynthetic pathway in-depth in order to increase thecontent of carotenoids in watermelon using transgenic techniques and carry outgermplasm innovation. The aim of establishing watermelon cotyledon tissue culturesystem and genetic transformation system is to provide technical support for thegenetic engineering of watermelon carotenoids and metabolic regulation. Effects ofcotyledon seedling age and different hormone concentrations on the induceddifferentiation of cotyledons and regeneration conditions watermelon cotyledons asthe explants were studied to establish efficient watermelon cotyledon tissue culturesystem．Then watermelon cotyledons as a receptor, effects of pre-culture time,Agrobacterium infection time, and co-culture time on confrontational buddifferentiation and effects of different antibiotic concentration on watermeloncotyledon growth were investigated to establish Agrobacterium-mediated PSY genetictransformation system．The following results were obtained：（1）The establishment of awatermelon cotyledon explant culture systemTheappropriate culture conditions for watermelon cotyledon tissue culture were obtainedthrough the experiments on several key factors. The appropriate time for cotyledonseedling age was4days, and time shorter or longer of cotyledon seedling age wasunfavorable for adventitious bud differentiation. After the cotyledons were culturedfor a certain time under dark the differentiation rate was high. The appropriate culturemedium was MS medium supplemented with2.0mg/L6-BA and0.5mg/L IBA,and MS medium supplemented with2.0mg/L6-BA and0.5mg/L IAA．（2）The establishment of Agrobacterium-mediated PSY genetic transformation system of watermelon cotyledon．Effects of different infection time (15,30,45,60min), pre-incubation time (2,3days)，co-culture time (2，3，4days) on confrontationalbud differentiation and effects of antibiotics (Kan，Amp，Cef) on the growth of ofwatermelon cotyledons were also studied cotyledons with4days seedling age asacceptor. The results indicated that75mg/L of Kan can be well used to screenresistance buds. Within two weeks200mg/L of Amp and400mg/L of Cef caninhibit largely Agrobacterium proliferation.2Days pre-incubation time wasappropriate. After15min of the bacteria infection the cotyledons were planted on MSmedium supplemented with2.0mg/L6-BA,0.5mg/L IBA,75mg/L Kan, and200mg/L Amp and cultured under dark conditions for3days, then transplanted onMS medium supplemented with2.0mg/L6-BA,0.5mg/L IBA,75mg/L Kan,and200mg/L Amp and cultured at a temperature of25℃±2℃under lightconditions of2000lx12h/d conditions for about30days. Resistant buds were wellobtained．（3）Four resistant plants were obtained through a series of experiments. Resistantplants examined by PCR methods were positive plants and the other three are falsepositive plants. The results showed that plant exogenous PSY was integrated into thewatermelon genome．...