Efficient plant regeneration and Agrobacterium-mediated transformation of Begonia semperflorens-cultorum

Begonia semperflorens-cultorum, known as wax begonia, is one of the most popular Begonia species in which variable commercial cultivars have been produced. The genetic transformation technique is required for further modification of this species because introduction of some desired traits such as novel flower colors cannot be achieved by conventional breeding. Here we report the procedures of an efficient plant regeneration from leaf segment and production of transgenic plants using Begonia semperflorens-cultorum. Efficient induction of adventitious shoots was achieved when the explants were cultured on MS medium supplemented with 1.5 mg/l thidiazuron (TDZ), 0.5 mg/l α-naphthaleneacetic acid (NAA), 30 g/l sucrose and 2.5 mg/l gellan gum. Agrobacterium tumefaciens strain EHA101 containing the plasmid pIG121-Hm was used for gene transfer. The transient GUS expression was significantly increased when 10 mM MES was added to the co-cultivation medium. By culturing the infected explants under the selection pressure with 10 mg/l hygromycin (Hm), the Hm-resistant independent shoots were obtained at a frequency of 0.78 per explant. The regenerated Hm-resistant plants showed the integration of hygromycin phosphotransferase (HPT) gene into the genome and stable GUS expression, as the proof of genetic transformation. Consequently, the conditions established in this study enabled the transformation of Begonia semperflorens-cultorum.