Enhanced Induction of Vir Genes Results in the Improvement of Agrobacterium - Mediated Transformation of Eggplant

Abstract

Induction of Agrobacterium vir genes is one of the basic requirements for T-DNA transfer and integration into plant genome. Here we study the vir gene induction by various explant types of eggplant in order to develop a transformation protocol with improved efficiency using binary vector constructs - harbouring a hygromycin phosphotransferase gene (hpt) as a selection marker and a gfp:gus fusion gene as a reporter. A protocol for efficient Agrobacterium-mediated transformation of eggplant (Solanum melongena L cv Pusa Purple Long) has been developed by optimizing factors. Leaf, cotyledon and hypocotyl explants were tested for their ability to induce Agrobacterium vir-genes using a VirE:lacZ fusion construct and were shown to be poor inducers of the same. Addition of 100 µM acetosyringone during infection and co-cultivation steps of transformation could enhance the vir gene induction as well as a 2–3 fold increase in transformation frequency. Transformed explants showed the expression of reporter genes gus and gfp. The transgenics were analysed by peR and Southern blot hybridization, and were shown to have T-DNA integrated into their genome. The data suggest that eggplant is a relatively poor inducer of Agrobacterium vir genes, probably due to minimal phenolic production, and by modulating vir gene induction using phenolics like acetosyringone eggplant transformation can be improved.