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

Abstract

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 β-glucuronidase (GUS) gene (uidA) as a scorable marker and a neomycin phosphotransferase gene (nptII) as a selectable marker. The seeds of agro-infiltrated plants (T0) were germinated on a medium containing 130 mg l−1 kanamycin, and the seedlings that remained green were considered T1 transgenic plants. Histochemical GUS assays, PCR, Southern analysis, and RT-PCR confirmed that both transgenes were integrated into the genome of T1 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.