Date of Award
Master of Science (MS)
Testing gene expression patterns is an important process in learning more about an organism. The standard methods of rtPCR and RNASeq provide highly detailed data on specific expression patterns, but can be resource consuming if many different samples including genes or organisms must be tested. A potential alternative for studying expression is the use of a reporter system carried by a vector system that encodes fluorescent proteins under the control of promoters of interest. Many lepidopterans (moths) are pestiferous and better understanding of gene expression levels in lepidopterans, both endogenous and exogenous including from their viruses, would be beneficial. Baculoviruses have been used widely as vectors to deliver genetic elements to lepidopteran hosts. Here, I developed a first generation baculovirus reporter vector to analyze gene expression regulatory elements in lepidopterans. I generated backbone transfer plasmids and viruses, including recombinant experimental viruses incorporating previously examined polydnavirus promoter elements. Recombinant viruses were used to infect Sf9 insect cells and fluorescence emission was measured by plate reader at several multiplicities of infection and times post-infection to estimate promoter activity. Infected cells were also observed via microscopy. I observed repeatable emission patterns associated with the inserted promoter elements, while identifying possible issues associated with the chosen reporter system that may be addressed in future modifications.
Howard, Daniel, "Development of a Novel Fluorescent Reporter Baculovirus Vector for Testing Promoters in Lepidopterans" (2023). All Theses. 4041.