Agrobacterium tumefaciens – Mediated transformation of Woodfordia fruticosa (L.) Kurz

Abstract

In the present study, a protocol for Agrobacterium tumefaciens-mediated transformation has been optimized for Woodfordia fruticosa (L.) Kurz. Precultured axenic leaf segments were cocultivated with A. tumefaciens strain LBA4404 harboring the binary plasmid pCAMBIA1301 with b-glucuronidase (uidA) containing intron as the reporter gene and hygromycin phosphotransferase (hpt) as a selectable marker gene. After 3 days of co-cultivation, leaf segments were cultured on MS medium containing Thidiazuron (TDZ 4.54 lM) and Indole-3-acetic acid IAA (1.14 lM) + 20 mg/ l hygromycin + 200 mg/l cefotaxime (PTSM 1 ) for 4 weeks (includes a single subculture onto the same medium at a 2 week interval). They were subsequently cultured for 3 weeks on MS medium containing Thidiazuron (TDZ 4.54 lM) and Indole-3-acetic acid IAA (1.14 lM) + 25 mg/l hygromycin + 100 mg/l cefotaxime (PTSM 2 ) medium for further development and shoot elongation. The hygromycin resistant shoots were rooted on a rooting medium (PTRM) containing half strength MS medium + 4.90 lM IBA + 25 mg/l hygromycin. A highest transformation efficiency of 44.5% with a mean number of 2.6 transgenic shoots per explant was achieved. Successful transformation was confirmed by the histochemical GUS activity of the regenerated shoots, PCR and RT-PCR analysis using respective primers. Southern blot analysis revealed that the hpt gene integrated into the genome of transgenic W. fruticosa. Establishment of genetic transformation protocol may facilitate the improvement of this medicinal plant in terms of enhancement of secondary metabolites.Ó 2015 Production and hosting by Elsevier B.V. on behalf

Agrobacterium tumefaciens-mediated transformation of Dendrobium lasianthera J.J.Sm: An important medicinal orchid

“…The insertion of foreign genes into the genome of plants mediated by Agrobacterium tumefaciens is an effective and reproducible method and has been successfully applied to various plants such as Artemisia carvifolia [5], Woodfordia fruticosa [6], Solanum trilobatum [7], Withania somnifera [8], Vanda kasem's [9], and Erycina pusilla [10].…”

Agrobacterium tumefaciens mediated transformation of chicory (Cichorium intybus L.) with Mytilus galloprovincialis foot protein type five (MGFP-5)

“…For the genetic transformation of plants, the preculture of leaf explants can promote cell division, which helps Agrobacterium infection and aid the integration of exogenous DNA, thus enhancing transformation efficiency (Birch 1997, Bullea et al, 2015. This was likely due to the likeliness of T-strand integration into cells that actively underwent cell cycle progression (Villemont et al, 1997).…”