Luciferase-mediated assay to detect the PAMP-triggered gene expression in transgenic Nicotiana benthamiana.

Luciferase is one of the bioluminescence-producing agents, which was widely used as a reporter enzyme for constructing bioassay systems to study gene expression with high accuracy and within a broad dynamic spectrum. Perception of pathogen associated molecular patterns (PAMPs) in plants often lead to significant transcriptional changes. The transcriptional changes of defense-related genes are often used as a marker to assay PAMP-triggered plant immune response. In this study, we showed that the marker gene CYP71D20 was rapidly activated in Nicotiana benthamiana upon treatment with the bacterial PAMP flg22 and the Phytophthora elicitin INF1. In addition, we generated transgenic N. benthamiana using the luciferase as a reporter gene to analyze CYP71D20 gene expression upon PAMP treatment. The transgenic line carrying the luciferase gene driven by CYP71D20 promoter was treated with the bacterial PAMP flg22 or Phytophthora elicitin INF1. Transcriptional activation of CYP71D20 was measured by monitoring the luciferase activity. The results showed that the LUC activity was increased after treatment with different PAMPs, indicating that the CYP71D20-promotor luciferase assay can be used to study the PAMP-triggered gene expression in N. benthamiana.