A Dual Luciferase Reporter System for Measures Transcriptional Activity during Tick-Pathogen Interactions.

Knowledge of the transcriptional responses of vector-borne pathogens at the vector-pathogen interface is critical for understanding disease transmission. () , the causative agent of Lyme disease in the United States, is transmitted by the bite of infected . ticks. It is known that has altered patterns of gene expression during tick acquisition, persistence and transmission. Recently, we and others have discovered expression of RNAs found internal, overlapping, and antisense to annotated open reading frames in the genome. However, there is a lack of molecular genetic tools for for quantitative, strand-specific, comparative analysis of these transcripts in distinct environments such as the arthropod vector. To address this need, we have developed a dual luciferase reporter system to quantify promoter activities in a strand-specific manner. We demonstrate that constitutive expression of a codon-optimized luciferase gene ( ) allows normalization of the activity of a promoter of interest when fused to the codon-optimized luciferase gene ( on the same plasmid. Using the well characterized, differentially regulated, promoters for flagellin (), outer surface protein A () and outer surface protein C (), we document the efficacy of the dual luciferase system for quantitation of promoter activities during growth and in infected ticks. Cumulatively, the dual luciferase method outlined herein is the first dual reporter system for , providing a novel and highly versatile approach for strand-specific molecular genetic analyses.