The Functional Identification Of Eugenol Synthetase From Rosa Chinensis ’Old Blush’

With the sustainable improvement of life quality, people have the high requirement about plant quality, who not only are satisfied the rich colour, also hope to have pleasant fragrance. The traditional breeding technology is very limited in floral improvement. For the sake of break this bottleneck and further improve the quality of the flowers, a large amount of floral genes have been cloned. So the genetic transformation system is becoming more and more important and urgent.In order to analyze the function of eugenol synthase gene from Rosa chinensis’old blush’, the embryo cells from’old blush’ are as a receptor for genetic transformation of genes. The woody plant genetic transformation need to overcome a lot of problem, so the genetic transformation’s experiment is also carried out on the model plant petunia. Meanwhile, RcEGS was transformed to Petunia hybrida. By detecting positive petunias’(Petunia hybrida) floral composition to the preliminary analysis the function of RcEGS in plants. These results are as follows:(1) Germination effect is better by6-BA1mg/L about Chinese rose’old blush’ axillary bud。The experiment is found that’old blush’ espitose buds is better on the rooting medium proliferation (1/2MS+NAA0.05mg/L), blade natural stretching, and it can propagate on proliferation medium, while leaves curled is bad for use later research;(2) The most of callus is induced from petiole, it is found that’old blush’has the highest callus induction at rate of90.33%nd has the best callus status, when the2,4-D is3mg/L; carbon source is glucose; explant is tissue culture seedlings. In additional, the ratio is up to95.00%, when AgNO3is10mg/L;(3) It is found that adventitious bud from’old blush’ ould be not induced by TDZ≥5mg/L, and it is gotten on the callus induction medium. The ratio of induction is0.21%;(4) According to the agrobacterium (OD600=0.5) containing the GUS gene infects ’old blush’ allus, the experiment is found that the transient expression ratio is very hight level to93.3%y agrobacterium infecting;(5) Agrobacterium EHA105with RcEGS gene (OD600=0.5) infect’old blush’ embryonic callus for15min and get’old blush’ eedlings for three months continuous culture; (6) Petunia hybrida is a model plant to verify for the function of scent-related gene, which contains respective substrate coniferyl alcohol. Using "semal method" to obtain petunia’ transgenic buds, they are screening through antibiotics for a positive plants and the level of DNA. The statistics found the success rate of petunias transforming is55%(7) By RT-PCR, it is found that RcEGS expression level is different in different strains of positive plants. The highest expression line is selected to test the scent. The petunia flowers’ volatiles were detected by GC-MS. The results are found the wild petunias do not contain methyl eugenol and transgenic petunias’ fragrance ingredients contain methyl eugenol which account for0.47%in the major components. In conclusion, RcEGS can regulate the synthesis of methyl eugenol.