High sensitivity and low-cost flavin luciferase (FLUX<sup>Vc</sup>)-based reporter gene for mammalian cell expression
Issued Date
2023-05-01
Resource Type
ISSN
00219258
eISSN
1083351X
Scopus ID
2-s2.0-85153360371
Pubmed ID
36965614
Journal Title
Journal of Biological Chemistry
Volume
299
Issue
5
Rights Holder(s)
SCOPUS
Bibliographic Citation
Journal of Biological Chemistry Vol.299 No.5 (2023)
Suggested Citation
Phonbuppha J., Tinikul R., Ohmiya Y., Chaiyen P. High sensitivity and low-cost flavin luciferase (FLUX<sup>Vc</sup>)-based reporter gene for mammalian cell expression. Journal of Biological Chemistry Vol.299 No.5 (2023). doi:10.1016/j.jbc.2023.104639 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/81594
Title
High sensitivity and low-cost flavin luciferase (FLUX<sup>Vc</sup>)-based reporter gene for mammalian cell expression
Author(s)
Other Contributor(s)
Abstract
Luciferase-based gene reporters generating bioluminescence signals are important tools for biomedical research. Amongst the luciferases, flavin-dependent enzymes use the most economical chemicals. However, their applications in mammalian cells are limited due to their low signals compared to other systems. Here, we constructed Flavin Luciferase from Vibrio campbellii (Vc) for Mammalian Cell Expression (FLUXVc) by engineering luciferase from V. campbellii (the most thermostable bacterial luciferase reported to date) and optimizing its expression and reporter assays in mammalian cells which can improve the bioluminescence light output by >400-fold as compared to the nonengineered version. We found that the FLUXVc reporter gene can be overexpressed in various cell lines and showed outstanding signal-to-background in HepG2 cells, significantly higher than that of firefly luciferase (Fluc). The combined use of FLUXVc/Fluc as target/control vectors gave the most stable signals, better than the standard set of Fluc(target)/Rluc(control). We also demonstrated that FLUXVc can be used for testing inhibitors of the NF-κB signaling pathway. Collectively, our results provide an optimized method for using the more economical flavin-dependent luciferase in mammalian cells.