An Agrobacterium-mediated transformation via organogenesis regeneration of a facultative CAM plant, the common ice plant Mesembryanthemum crystallinum L

Abstract

The common ice plant, Mesembryanthemum crystallinum L. provides a useful model for the study of environmentally induced photosynthetic conversion and abiotic stresses tolerance. However, a procedure for the production of transgenic ice plant, which is essential for functional genomics, has not been fully established. Here we tested the factors on the transformation of cotyledonary nodes excised from the ice plant seedlings such as thidiazuron (TDZ), NaCl and phytosulfokine (PSK), a peptidyl plant growth factor using Agrobacterium tumefaciens strains EHA101 and EHA105 harboring binary vector plasmids pBI7EGFP and pCAMBIA1302, respectively. The established procedure is as follows: the explants (cotyledonary nodes) were co-cultivated with Agrobacterium for 3 days, and the explants were cultured in the medium with 0.5 mg l⁻¹ kinetin and 100 mg l⁻¹ carbenicillin for 72 h, and they were cultured in the medium with 0.5 mg l⁻¹ kinetin and 100 nM PSKfor 4 weeks. Thidiazuron and NaCl enhanced the production of multiple adventitious shoot formation during regeneration but reduced the transformation efficiency due to the vitrification of adventitious shoots. PSK was effective in the production of healthy adventitious shoots. The transformation frequency at the stage of whole plants was 0.6% and 4.6% per inoculated cotyledonary nodes using the Agrobacterium strain EHA101 (pBI7EGFP) and EHA105 (pCAMBIA1302), respectively.