Genetic transformation of an Ac/Ds-based transposon tagging system in carrot (Daucus carota L.)

Abstract

Maize transposable elements, Activator (Ac) and Dissociation (Ds) have been transformed into several heterologous plant species and they have been successfully used to tag and clone genes in Arabidopsis, flax, petunia, tobacco, and tomato. To investigate the possibility of transposon tagging for subsequent cloning of carrot genes, carrot calli were transformed with a modified maize transposable element Activator-transposase (Ac-TPase) and Ds using an Agrobacterium-mediated transformation method. Ds excision was observed in the tissue culture-regenerated plants carrying both Ac-TPase and Ds. Analysis of Ds footprints in six plants demonstrated the occurrence of small deletions at the donor site. Reinsertion of Ds into new chromosomal sites was detected in the transgenic plants analysed and the analysis of sequences flanking Ds at the insertion sites using TAIL-PCR demonstrated that 37.5% of Ds insertions were into gene regions in the carrot genome. Although previous studies suggested that the autonomous Ac element may not be a suitable transposon for insertional mutagenesis in carrot, our results deduced that the Ac/Ds based two-element transposon tagging system can be another tool to mutagenise the carrot genome for the purpose of cloning carrot genes.