A quantitative autonomous bioluminescence reporter system with a wide dynamic range for Plant Synthetic Biology

Abstract

ABSTRACTEngineered autonomous bioluminescence (EAB) offers many potential applications in Plant Synthetic Biology, notably asin vivoreporter system. Current EAB reporter configurations are limited for quantitative applications due to low dynamic range. We reconfigured theNeonothopanus nambifungal bioluminescence (NeoLuc) pathway to serve as a high-throughput and inexpensive reporter for quantitative analysis of gene expression. We showed that by configuring the first committed step in the pathway (HispS) as the transcriptional entry point instead of the fungal luciferase, the dynamic range of the output increased dramatically, equaling that of the FLuc/RLuc reporter, and outperforming it in high throughput capacity. Furthermore, the inclusion of an enhanced GFP as normalizer allowed transient ratiometric measurements inN. benthamiana. Fast and rich datasets generated by the NeoLuc/eGFP system enabled us to undertake the optimization of new challenging synthetic gene circuits, including a complex agrochemical/optogenetic dual input switch for tight control of engineered metabolic pathways.