The floral inoculating protocol: A simplified Arabidopsis thaliana transformation method modified from floral dipping

Abstract

The floral dip protocol mediated by Agrobacterium tumefaciens is the most widely used transformation method for Arabidopsis thaliana. The "floral dip" process in which A. thaliana flower buds are dipped in an Agrobacterium cell suspension requires large volumes of bacterial cultures grown in liquid media, large shakers and centrifuges, and experimental space for them. These factors limit the number of transformations that can occur at once. We established that A. thaliana can be transformed by inoculating 5m l of Agrobacterium cell suspension in flower buds, thus avoiding the use of large volumes of Agrobacterium culture. Using this modified protocol, we obtained 15-50 transgenic plants per transformation from each pot containing 3 A. thaliana plants. The protocol is satisfactory to be used for subsequent analyses. This simplified method, without floral dipping, which requires large volumes of Agrobacterim culture, offers as efficient a transformation as previously reported protocols. This method reduces the required workload, cost, time, and space. Furthermore, an important aspect of this modified protocol is that it allows many independent transformations to be performed at once. © 2010 The Japanese Society for Plant Cell and Molecular Biology.