Genetic transformation of Ginkgo biloba by Agrobacterium tumefaciens

A reproducible protocol has been established for the transformation of Gink go biloba by Agrobacterium tumefaciens. Embryos were co-cultivated with Agr obacterium tumefaciens GV3101 (pGV2260) carrying the binary vector pTHW136, which contained the gus reporter gene and the nptII selectable gene, encod ing the enzymes beta-glucuronidase (GUS) and neomycin phophotransferase II, respectively. Transient GUS activity has been used to screen the Effects o f different factors on the transfer of UNA into embryos (age of embryos, in fection method, composition of co-cultivation medium). Then, experimental c onditions have been defined to obtain transgenic kanamycin-resistant G, bil oba calluses expressing GUS activity, The highest rate of transformation (4 5%) was reached using 1.5-month-old embryos co-cultivated on a medium lacki ng mineral elements. The integration of gas and nptII genes in calluses mas confirmed by polymerase chain reaction analysis and Southern blot analysis .