BACTERIAL LUCIFERASE PRODUCED WITH RAPID-SCREENING BACULOVIRUS VECTORS IS A SENSITIVE REPORTER FOR INFECTION OF INSECT CELLS AND LARVAE

Bacterial luciferase, derived from a fusion of the luxA and luxB genes of Vibrio harveyi, has been expressed at very high levels in caterpil lars and insect cells. The coding sequence for luciferase was inserted into vectors developed in our laboratory which were designed to exped ite screening of recombinant virus. These vectors contained the beta-g alactosidase indicator gene under control of immediate early (IE1), ea rly (ETL), or very late (P10) promoters and a cloning site for inserti ng the fused luciferase gene next to the polyhedrin promoter. Recombin ant baculoviruses containing the luciferase gene as well as the beta-g alactosidase gene could be easily selected when Bluo-gal (beta-galacto sidase indicator) was included in the plaque assays. Using cells deriv ed from the fall armyworm (Spodoptera frugiperda), luciferase was stro ngly expressed very late in infection (48-72 h). The bacterial lucifer ase assay was sufficiently sensitive that light production could be de tected from an extract of a single cell. In addition, live insects, in cluding the cabbage looper (Trichoplusia ni) and saltmarsh caterpillar (Estigmene acrea) were infected by mixing recombinant baculovirus int o their diet. Cabbage loopers (with an average wet weight of 223 mg) p roduced at least 195 mug of active luciferase and levels of synthesis peaked between 96-120 h. The results indicate that bacterial luciferas e may be used as a reporter of gene expression in insects.