Temporal characterization of protein production levels from baculovirus vectors coding for GFP and RFP genes under non-conventional promoter control

ABSTRACT The ease of use and versatility of the Baculovirus Expression Vector System (BEVS) has made it one of the most widely used systems for recombinant protein production However, co‐expression systems currently in use mainly make use of the very strong very late p10 and polyhedron (polh) promoters to drive expression of foreign genes, which does not provide much scope for tailoring expression ratios within the cell. This work demonstrates the use of different Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) promoters to control the timing and expression of two easily traceable fluorescent proteins, the enhanced green fluorescent protein (eGFP), and a red fluorescent protein (DsRed2) in a BEVS co‐expression system. Our results show that gene expression levels can easily be controlled using this strategy, and also that modulating the expression level of one protein can influence the level of expression of the other protein within the system, thus confirming the concept of genes “competing” for limited cellular resources. Plots of “expression ratios” of the two model genes over time were obtained, and may be used in future work to tightly control timing and levels of foreign gene expression in an insect cell co‐expression system. Biotechnol. Bioeng. 2015;112: 1822–1831. © 2015 Wiley Periodicals, Inc. Scatterplot of red and green fluorescence intensity of two proteins over time in insect cells infected with a basic‐gfp‐polh‐rfp polycistronic baculovirus vector. These results show that co‐expression of foreign genes in insect cells can be finely controlled, and these genes compete with each other for access to resources for the production of their proteins.