BIOLISTIC TRANSFORMATION OF CHRYSANTHEMUM WITH THE NUCLEOCAPSID GENE OF TOMATO SPOTTED WILT VIRUS

In vitro regeneration and biolistic transformation procedures were dev eloped for several commercial chrysanthemum Dendranthema grandiflora T zvelev, syn. Chrysanthemum morifolium Ramat. cultivars using leaf and stem explants. Studies on the effect of several growth regulators and kanamycin on chrysanthemum regeneration were conducted, and a step-wis e procedure to optimize kanamycin selection and recovery of transgenic plants was developed. A population of putative transformed chrysanthe mum plants cvs. Blush, Dark Bronze Charm, Iridon, and Tara, was obtain ed after bombardment with tungsten microprojectiles coated with the bi nary plasmid pBIN19 containing the nucleocapsid (N) gene of tomato spo tted wilt virus (TSWV) and the marker gene neomycin phosphotransferase (NPT II). PCR analysis of 82 putative transgenic plants selected on k anamycin indicated that the majority of the lines (89%) were transform ed and contained both genes (71%). However, some transgenic lines cont ained only one of the genes: either the NPT II (15%) or the TSWV (N) g ene (14%). Southern blot analysis on selected transgenic lines confirm ed the integration of the TSWV (N) gene into the chrysanthemum genome. These results demonstrate the development of an efficient procedure t o transfer genetic material into the chrysanthemum genome and selectiv ely regenerate transgenic chrysanthemum plants at frequencies higher t han previously reported.