(127b) Accelerated Virus-Less Generation of Stable Insect Sf9 Cell Lines for High-Yield Production of Influenza Vaccines
AIChE Annual Meeting
Monday, October 29, 2018 - 12:48pm to 1:06pm
Taking advantage of the recently published Sf9 genome, we successfully employed bioinformatics tools and identified hypothetical genes responsible for nonhomologous end joining (NHEJ). More importantly, we confirmed their biological function, as their knockout via CRISPR resulted in stable Sf9 cell lines with improved efficiency in site-specific gene integration through homologous directed repair (HDR). These engineered Sf9 cell lines have enabled rapid generation of high-yield stable cell lines without the need for limiting dilution. Specifically, two Sf9 stable cells lines each producing high-yield influenza hemagglutinin subtype 1 (H1) and subtype 3 (H3) were generated within just one week. In addition to improving the efficiency of HDR for gene knockin, NHEJ-gene KO also improved the efficiency of using CRISPR to knockout other genes that are important for expressing human-like glycosylated H1/H3 proteins for vaccination. This system enables high-throughput production of recombinant HA to develop improved vaccines for each yearâs influenza season. Future work will focus on the production of more complex protein therapeutics such as multivalent subunit vaccines or complex virus-like particles. The accelerated generation of insect cell lines offers an attractive platform to advance the development and manufacture of any protein therapeutic.