The insect cell/baculovirus expression vector system (BEVS) is becoming increasingly popular for the production of recombinant proteins. The system offers a number of advantages, including:

  • Able to perform complex post-translational modifications (PTMs)
  • High success rate of soluble protein recovery
  • Suitable for the production of large protein complexes
  • High protein expression levels compared to other higher eukaryotes

The ability of insect cell/BEVS to generate proteins with complex PTMs, coupled with high expression levels, makes it particularly suitable for the production of mammalian proteins.

Insect cell/BEVS utilises the naturally occurring baculovirus, Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV), which infects insects from the Lepidopteran genus (moths and butterflies). Late in the infection cycle, the virus produces large amounts of polyhedral or inclusion bodies in the insect host, typically around 50% of the total cellular protein. This is exploited in the BEVS, where insect cells are infected with a modified AcMNPV carrying the recombinant gene of interest under control of the strong polyhedrin promoter (polh).

Insect Cell/BEVS Expression Systems

There are a number of different systems that can be used to generate a recombinant baculovirus containing the gene of interest. Some of the more common systems are listed below.

System Transfer Vectors Features
Bac-to-Bac pFastbac-1,
pFastBac Dual
Requires integration with baculovirus shuttle vector (Bacmid) in bacterial host cell before transfection
flashBAC pBac-1 Transfection with linear DNA, recombination occurs within host insect cells. Deletion of chiA enables better secretion of recombinant protein
BaculoGOLD pVL1392, 1393 Transfection with linear DNA, recombination occurs within host insect cells
BaculoDirect pENTR Transfer vector recombines with linear DNA before transfection into host cell. Requires Ganciclovir as selection agent

Cell Lines

The most common cell lines for use in the BEVS are Sf9 and High FiveTM. Both can be grown as adherent or suspension cultures and scaled up from microplates in a high throughput format to shake flasks and WAVE bioreactors. They are also able to be grown in serum-free media (SFM) and do not require the additon of CO2 gas.

Some commonly used insect cell lines are listed below.

Cell Line Origin Features
Sf9 Spodoptera frugiperda Adapted to SFM, commonly used to express all types of recombinant proteins
Mimic Sf9 Spodoptera frugiperda Adapted to SFM, expresses mammalian glycosyltransferases to generate proteins with more complex N-glycan structures
High FiveTM Trichoplusia ni Adapted to SFM, commonly used for secretion of recombinant proteins