Genetic transformation of<i>Ginkgo biloba</i>by<i>Agrobacterium tumefaciens</i>

Abstract

A reproducible protocol has been established for the transformation of Ginkgo biloba by Agrobacterium tumefaciens . Embryos were co‐cultivated with Agrobacterium tumefaciens GV3101 (pGV2260) carrying the binary vector pTHW136, which contained the gus reporter gene and the nptII selectable gene, encoding the enzymes β ‐glucuronidase (GUS) and neomycin phophotransferase II, respectively. Transient GUS activity has been used to screen the effects of different factors on the transfer of DNA into embryos (age of embryos, infection method, composition of co‐cultivation medium). Then, experimental conditions have been defined to obtain transgenic kanamycin‐resistant G. biloba calluses expressing GUS activity. The highest rate of transformation (45%) was reached using 1.5‐month‐old embryos co‐cultivated on a medium lacking mineral elements. The integration of gus and nptII genes in calluses was confirmed by polymerase chain reaction analysis and Southern blot analysis.