A non-tissue culture approach for developing transgenic Brassica juncea L. plants with Agrobacterium tumefaciens (original) (raw)

A non-tissue culture approach for the generation of transgenic Indian mustard (Brassica juncea) plants using Agrobacterium tumefaciens was developed. Inflorescences with floral buds were vacuum infiltrated with a suspension of A. tumefaciens strain EHA105 carrying a binary vector with an intron-containing βglucuronidase (GUS) gene (uidA) as a scorable marker and a neomycin phosphotransferase gene (nptII) as a selectable marker. The seeds of agro-infiltrated plants (T 0 ) were germinated on a medium containing 130 mg l −1 kanamycin, and the seedlings that remained green were considered T 1 transgenic plants. Histochemical GUS assays, PCR, Southern analysis, and RT-PCR confirmed that both transgenes were integrated into the genome of T 1 plants and were stably transmitted and expressed for over three generations. The transformants were obtained within 3-4 mo at a transformation frequency of 0.8%. This method may facilitate functional genomics and improvement of Brassica with novel desirable traits and with less time and expense.