A Simplified Floral Dip Method for Transformation of Brassica napus and B. carinata

Abstract

A floral dip method of genetic transformation has been developed in two different species of Brassica namely B. napus cv Elect and i Pusa Gaurav. Binary vector containing B. carinata Zinc Finger transcription factor (BcZF1) gene under the transcriptional control of abiotic stress-inducible Late Embryogenesis Abundant 1 (i) promoter was transformed into Agrobacterium tumefaciens strain GV-3101. This strain was used for transformation of Brassica spp via Agrobacterium-mediated floral dip method. Agrobacterium cells cultured in Yeast Extract Mannitol (YEM) were harvested and resuspended in half MS solution containing 5% sucrose and 0.05% Silwet L-77. The transformation was performed by dipping inflorescence at the initial blossom stage. Seeds from these plants were harvested and screened by germinating them on MS medium supplemented with kanamycin (25 mg l⁻¹). The kanamycin resistant putative transgenic plants were further confirmed by PCR and RT PCR using nptll gene-specific primers. The results showed that the Agrobacterium-mediated floral-dip transformation can be a successful strategy to develop transgenic B. napus and B. carinata in a short time without undergoing cumbersome vacuum infiltration or tissue culture steps.