Benchmarking conven/onal and vacuum-assisted floral dip transforma/on: A compara/ve protocol

Floral-dip transformation is a simple and widely used approach for generating transgenic plants in Brassicaceae. However, transformation efficiency varies considerably among species and genotypes, and the optimal transformation strategy often remains unknown when working with non-model taxa. This protocol provides a practical benchmarking workflow for the initial evaluation of floral-dip transformation using two commonly employed approaches: conventional floral dip (FD) and vacuum-assisted floral dip (VAFD). The protocol describes the preparation of Agrobacterium tumefaciens cultures, plant treatment, post-transformation handling, and subsequent seed screening. By directly comparing transformation efficiencies obtained with FD and VAFD, researchers can rapidly identify the more suitable approach for a given species or genotype before undertaking further optimization of transformation parameters. The workflow is particularly useful for establishing transformation protocols in previously untransformed or recalcitrant Brassicaceae species and provides a foundation for subsequent fine-tuning of transformation conditions.