The flashBAC system maximises production of secreted and membrane-targeted proteins by deletion of the chitinase gene from the virus genome. Tests with a wide range of secreted and membrane targeted proteins indicate that this chitinase-deficient virus can increase the yield of protein by up to 60-fold (in comparison with recombinant viruses that synthesise chitinase) (see ref. 1) However, chitinase deletion has no effect on the yield of recombinant proteins targeted to the cytoplasmic or nuclear compartments.

Baculovirus genomes contain several auxillary genes, which are non-essential for replication in insect cell culture. One of these is chitinase (chiA), which encodes an enzyme with exo- and endochitinase activity (see ref 2). In an infected insect, chitinase (together with cathepsin) facilitates host cuticle breakdown and tissue liquefaction at the very late stages of infection, so releasing the virus to infect more hosts (see ref 3). This function is obviously not required in cell culture. Confocal and electron microscopy observations of insect cells infected with AcMNPV have shown that chitinase is targeted to the endoplasmic reticulum (ER) where it is densely packed in a para-crystalline array, severely compromising the function and efficacy of the secretory pathway (refs 4,5). Deletion of chiA from flashBAC has no effect on virus replication, production of recombinant virus or the function of the polyhedrin gene promotor (controlling foreign gene expression).

Reference Papers

1. Possee, R. D.; Saville, G. P.; Thomas, C. J.; Patminidi, A. & King, L. A. (2001).
Recent advances in the development of baculovirus expression vectors. In Prospects For The Development Of Insect Factories. Proceedings of a Joint International Symposium of Insect COE Research Program and Insect Factory Research Project. October 22-23, Tsukuba, Japan.

2. Hawtin, R. E., Arnold, K., Ayres, M. D., Zanotto, P. M. d. A., Howard, S. C., Gooday, G. W., Chappell, L. H., Kitts, P. A., King, L. A., and Possee, R. D. (1995).
Identification and Preliminary Characterization of a Chitinase Gene in the Autographa californica Nuclear Polyhedrosis Virus Genome. Virology 12, 673-685.

3. Hawtin, R. E., Thomas, C. A., Gooday, G. W., Kuzio, J. A., King, L. A. and Possee, R. D. (1997).
Liquefaction of Autographa californica nucleopolyhedrovirus-infected cells is dependent on the integrity of virus-encoded chitinase and cathepsin genes. Virology, 238, 243-253.

4. Thomas, C. A., Hawes, C. R., Lee, B. Y., Min, M-K, King, L. A. and Possee, R. D. (1998).
Localisation of a baculovirus-induced chitinase in the insect cell endoplasmic reticulum. J Virol. 72, 10207-10212.

5. Saville G. P., Patmanidi A. L., Possee R. D., King L. A. (2004).
Deletion of the Autographa californica nucleo-polyhedrovirus chitinase KDEL motif and in vitro and in vivo analysis of the modified virus. J Gen Virol. 85, 821-31.