L-methionine sulfoximine as a novel selection agent for genetic transformation of orchids

Abstract

A key challenge in molecular breeding of orchids is the creation of efficient and reproducible gene transformation systems. In this study, we report a new transformation method utilizing L-methionine sulfoximine (MSO) as a novel agent for selection of transgenic Dendrobium hybrids D. Madame Thong-In and D. Chao Praya Smile with the bialaphos resistance (bar) gene as a selectable marker. Gene transformation was performed by biolistic bombardment with a 4-day recovery period on MSO-free medium and two selection stages on media with increasing amounts of selection agent, using concentrations of 5 and 10 microM MSO for D. Madame Thong-In, and 0.5 and 2 microM MSO for D. Chao Praya Smile. Independent transgenic orchid lines were obtained and the presence of the transgene was confirmed by PCR and Southern blot analysis. Because of substantial time and economic savings, the new transformation system using MSO as a selection agent will facilitate functional studies on orchid genes and genetic engineering of orchids with commercially valuable traits.