Energy transfer between fluorescent proteins using a co-expression system in Mycobacterium smegmatis

The goal of this study was to establish a two-plasmid co-expression system for Mycobacterium smegmatis. Two vectors with compatible origins of replica tion and a polylinker, which allows modular cloning of promoters and genes, were constructed and used to clone genes encoding a blue fluorescent prote in (BFP) and a green fluorescent protein (GFP). A 160-fold variation of GFP expression levels in M. smegmatis was achieved by combining three promoter s with different copy numbers of the vectors. An efficient energy transfer between BFP and GFP in M. smegmatis was observed by fluorescence measuremen ts and demonstrated that these genes were simultaneously expressed from bot h vectors. Thus, these vectors will be valuable for all strategies where co -expression of proteins in M. smegmatis is needed, e.g. for constructing a two-hybrid system or for deleting essential genes. (C) 2001 Elsevier Scienc e B.V. All rights reserved.