Studies On Genetic Transformation Of Narcissus Tazetta L.var. Chinensis Roem. By IPT Gene

Studies on Genetic Transformation of Narcissus tazetta L.var. chinensis Roem. by IPT GeneABSTRACTNarcissus tazetta has great value as an important flower plant in China. The autolatory senescence inhibition system, constructed by fusing leaf senescence specific promoter with IPT gene, is applied to the transformation of narcissus in this experiment. JPT expression in the transgenic plant can delay leaf senescence by increasing endogerous cytokinin level. At the same time, we have in the first established genetic transformation system of narcissus through the guide of GUS reporter gene. The main results are as below:1.Tissue culture procedure in the narcissus genetic transformation was developed. All of bulbs initiated regeneration of bulbils directly when cultured in MS medium supplemented with 6-BA(4mg/L)+ NAA(O.5mgIL), the coefficient of regeneration is 11.2; At the other hand, about 74.2% of flower peduncles induced calli when cultured in MS+ 6-BA(lmg/L)+NAA(5mgIL), and the calli can differentiate great many bulbils in MS+6-BA(4mg/L)-T- NAA(0.Smg/L), the coefficient of differentiation is 12.9.2.Established efficient transgenic technology of narcissus. The chimeric PC2MV35S-GUS and PSAG)2-IPT genes were transformed into narcissus mediated by Agrobacterium tumefaciens. About 95% of bulbs indicated expression of GUS reporter gene after co-cultivating 3 days with agrobacterium. so did 85% of flower peduncles. Otherwise, there were 15% of transformed bulbs and all of flower peduncles that could regenerate bulbils.3.Interval-screening was adopted to screen transformed narcissus tissue.29.3% of transformed tissue obtained the resistance to Hyg. and 20% of the Hygr tissue indicated expression of GUS gene. Transformed tissue regenerated bulbils in MS+6-BA(4mgIL)+ NAA(0.5mgIL), the coefficient of regeneration is 3.4. There had expression of GUS gene in transgenic plant leaf.4.PCR analysis showed that 14% of the HygT tissue had positive band; PCR-Southern indicated that it was the fragment of PSAGI2. Southern analysis showed that the foreign gene had been intigrated into narcissus chromosome randomly.