The firefly luciferase gene as a non-invasive reporter for Dendrobium transformation

Abstract

Here a screening method is described for transformed tissues and transgenic plants of Dendrobium (Orchidaceae) using the firefly luciferase gene (luc) as a combined marker/reporter gene. Protocorm-like-bodies (PLB) were bombarded with tungsten particles (1.3 μm) coated with plasmids carrying a 35S-luc chimeric gene. Three weeks after bombardment 1 mM luciferin was added to the tissues and transformed cells were identified by virtue of their bioluminescence as monitored by low-light video microscopy in combination with a real-time photon imaging technique. Transformed tissues were excised, allowed to proliferate, and then subjected to a second round of screening. After three rounds of growth and screening, transformed Dendrobium tissues expressing luciferase were used to generate transgenic plants. Southern blot analysis of several transgenic lines confirmed the integration of the luciferase gene into the orchid genome. It is thought that this procedure can be used for transformation of not only orchids but other species as well.