Recovery of herbicide-resistant Azuki bean [Vigna angularis (Wild.), Ohwi & Ohashi] plants via Agrobacterium-mediated transformation

Abstract

Transgenic azuki bean [Vigna angularis (Willd.) Ohwi & Ohashi] plants expressing the hygromycin phosphotransferase (hpt), green fluorescent protein (sgfp) and phosphinothricin acetyltransferase (bar) genes were obtained by Agrobacterium- tumefacients - mediated transformation. A total of 210 epicotyl explants were inoculated with A. tumefaciens strain EHA105, harboring the binary plasmid pZHBG on MS co-cultivation medium supplemented with 100 mM acetosyringone and 10 mg/l of BA. Following selection on MS medium with 15 mg/l of hygromycin, the regenerated adventitious shoots that formed on the induced calli were further screened for sgfp expression before transferred to rooting medium. 31 transgenic plants were obtained with transformation frequency of 14%. The presence of transgenes in transformed azuki bean plants was confirmed by polymerase chain reaction (PCR) and southern blot analysis. Transcription of the bar and hpt genes was assessed by reverse transcription polymerase chain reaction (RT-PCR) analysis. sgfp- positive transgenic plants exhibited functional expression of the bar gene as determined by assaying for resistance to bialaphos applied directly to leaves. This result demonstrates the feasibility of introducing potentially useful agronomic traits into azuki bean through genetic engineering.

Key Words: Agrobacterium tumefaciens, bar gene, bialaphos, transgenic, Vigna angulazris.

African Journal of Biotechnology Vol.4(1) 2005: 61-67