Autonomous Bioluminescent Expression of the Bacterial Luciferase Gene Cassette (lux) in a Mammalian Cell Line
Open Access
- 27 August 2010
- journal article
- research article
- Published by Public Library of Science (PLoS) in PLOS ONE
- Vol. 5 (8), e12441
- https://doi.org/10.1371/journal.pone.0012441
Abstract
The bacterial luciferase (lux) gene cassette consists of five genes (luxCDABE) whose protein products synergistically generate bioluminescent light signals exclusive of supplementary substrate additions or exogenous manipulations. Historically expressible only in prokaryotes, the lux operon was re-synthesized through a process of multi-bicistronic, codon-optimization to demonstrate for the first time self-directed bioluminescence emission in a mammalian HEK293 cell line in vitro and in vivo. Autonomous in vitro light production was shown to be 12-fold greater than the observable background associated with untransfected control cells. The availability of reduced riboflavin phosphate (FMNH2) was identified as the limiting bioluminescence substrate in the mammalian cell environment even after the addition of a constitutively expressed flavin reductase gene (frp) from Vibrio harveyi. FMNH2 supplementation led to a 151-fold increase in bioluminescence in cells expressing mammalian codon-optimized luxCDE and frp genes. When injected subcutaneously into nude mice, in vivo optical imaging permitted near instantaneous light detection that persisted independently for the 60 min length of the assay with negligible background. The speed, longevity, and self-sufficiency of lux expression in the mammalian cellular environment provides a viable and powerful alternative for real-time target visualization not currently offered by existing bioluminescent and fluorescent imaging technologies.Keywords
This publication has 32 references indexed in Scilit:
- Non-Invasive Detection of a Small Number of Bioluminescent Cancer Cells In VivoPLOS ONE, 2010
- Mathematical model of the Lux luminescence system in the terrestrial bacterium Photorhabdus luminescensMolecular BioSystems, 2008
- Codon optimization of bacterial luciferase (lux) for expression in mammalian cellsJournal of Industrial Microbiology & Biotechnology, 2005
- Whole-body fluorescence lifetime imaging of a tumor-targeted near-infrared molecular probe in miceJournal of Biomedical Optics, 2005
- Advances in In Vivo Bioluminescence Imaging of Gene ExpressionAnnual Review of Biomedical Engineering, 2002
- Toxicity of the bacterial luciferase substrate, n‐decyl aldehyde, to Saccharomyces cerevisiae and Caenorhabditis elegansFEBS Letters, 2001
- THE GREEN FLUORESCENT PROTEINAnnual Review of Biochemistry, 1998
- Fusion of luxA and luxB and its expression in E. coli, S. cerevisiae and D. melanogasterJournal of Bioluminescence and Chemiluminescence, 1990
- Induction of fatty aldehyde dehydrogenase activity during the development of bioluminescence in Beneckea harveyiBiochemical and Biophysical Research Communications, 1976
- Conversion of aldehyde to acid in the bacterial bioluminescent reactionBiochemistry, 1973