Baculovirus production

Batista FRX, Pereira CA, Mendon a RZ, Moraes AM (2005), Enhancement of Sf9 cells and baculovirus production employing Grace s medium supplemented with milk whey ultrafiltrate, Cytotechnology 49 1-9. 

This chapter describes the application of animal cells, particularly insect cells, for baculovirus production and its use as a bioinsecticide. The use of baculovirus in the control of insect pests and its production in vitro are discussed. 

In vitro baculovirus production can also be affected by the cell line. Polyhedra production yield can vary between different tissues within the same host as well as in different cell lines. Furthermore, the cell line itself may be a source of viral instability in the form of transposon-mediated mutagenesis (Fraser et al., 1985). (Transposons are sequences of DNA that can move around to different positions within the genome, a process called transposition. In the process, they can cause mutations, and change the amount of DNA in the genome.)... 

Andersen, J.N., "Temperature Effect on Recombinant Protein Production Using a Baculovirus/Insect Cell Expression System", Diploma Thesis, University of Calgary and Technical University of Denmark, 1995. 

Jarvis, D.L. (2003) Developing baculovirus-insect cell expression systems for humanized recombinant glycoprotein production. Virology, 310 (1), 1-7. 

Lamberg, A., Helaakoski, T., Myllyharju, J., Peltonen, S., Notbohm, H., Pihlajaniemi, T. and Kivirikko, K.I. (1996) Characterization of human type III collagen expressed in a baculovirus system - production of a protein with a stable triple helix requires coexpression with the two types of recombinant prolyl 4-hydroxylase subunit.Journal of Biological Chemistry 271, 11988-11995. 

Baculovirus Improved procedure Infection of insect cells High expression yields Relatively slow virus production Different post-translational processing... 

Insect cells in culture are also hosts for recombinant protein production. Production of recombinant proteins in the baculovirus expression vector system is the most common system. Titers of recombinant protein as high as 11 g/L have been obtained. 

Insect cell expression Baculovirus High-level expression Formation of disulphides Glycosylation Glycosylated product may differ from mammalian systems Not necessarily fully functional... 

Smith, G. E., Summers, M. D., and Fraser, M. J. (1983). Production of human beta interferon in insect cells infected with a baculovirus expression vector. Mol. Cell Biol. 3, 2156-2165. 

For other production hosts (yeast, insect, and mammalian cells), standard promoter formats have been used in combination with FITP cloning methods to produce vectors for expression screening (see Section 2.3.2). A particularly interesting development is the use of multipromoter plasmids for expression in two or more hosts from a single vector. The construction of a dual E.coli (T7 promoter) and baculovirus transfer vector (polH promoter) for expression in insect cells has been described (Chambers et al., 2004). A three-promoter vector (T7, plO, and hCMV or CAG promoter) is available from Novagen (pTrlEX ) and its use reported for comparing protein expression in E. coli and insect cells (Xu and Jones, 2004). 

Production of Core and Virus-Like Particles with Baculovirus Infected Insect Cells... 

In this paper the fundamental aspects of process development for the production of core and virus-like particles with baculovirus infected insect cells are reviewed. The issues addressed include particle formation and monomer composition, chemical and physical conditions for optimal cell growth, baculovirus replication and product expression, multiplicity of infection strategy, and scale-up of the process. Study of the differences in the metabolic requirements of infected and non-infected cells is necessary for high cell density processes. In the bioreactor, the specific oxygen uptake rate (OURsp) plays a central role in process scale-up, leading to the specification of the bioreactor operational parameters. Shear stress can also be an important variable for bioreactor operation due to its influence on cell growth and product expression. 

Multi-gene Baculovirus Vectors and Chimeric Particle Production. . 190... 

The Baculoviridae are a family of large enveloped DNA viruses that are characterised by rod-shaped nucleocapsids and relatively large double stranded DNA genomes. Autographa californica Multicapsid Nuclear Polyhedrosis Virus (AcMNPV) is the baculovirus most currently used as vector for protein production with insect cells. Several reviews are available describing baculovirus structure and its molecular biology [6-8]. 

The industrial application of CLPs and VLPs is in the development phase. This can be expected since the baculovirus-insect cells system has become one of the most popular systems for heterologous protein and CLP/VLP production at laboratory scale. After defining the appropriate particle composition for the viruses of interest, research is now addressing the engineering issues in this system (Scheme 1). 

In this paper the fundamental aspects concerning the production of CLPs and VLPs with baculovirus infected insect cells are reviewed. It is not the goal of this communication to review all the aspects of insect cell culture technology, since this would be a task impossible to achieve in a single chapter. The interested reader should refer to the references. This review is structured in four parts ... 

In this part we deal with the major chemical and physical factors affecting insect cell growth, baculovirus replication and product expression. The issues addressed are cell line selection for product expression, baculovirus-cell interactions and impact of differences in metabolic requirements of infected and non-infected cells in overall productivity. 

In this part the application of mathematical models to CLP and VLP production with baculovirus infected insect cell cultures is discussed. Special emphasis on model evaluation is made along with the definition of directions in future process development research with this system. 

For the production of CLPs and VLPs with baculovirus infected insect cells the specific proteins that are required for particle formation should be chosen at an early step. The specific particle composition, along with the expression of other non-structural (NS) proteins often has a great impact on particle stability [11] or on particle localisation [12], i.e. - cell-associated or secreted to the supernatant. 

Different approaches have been reported in the Hterature regarding the number of genes cloned in the same baculovirus. Pioneering work has been done at Prof Polly Roy s laboratory at Oxford University for Bluetongue virus (BTV) CLP and VLP production, where these authors generated BTV-CLP and BTV-VLP of different types and monomer compositions [4,13-16]. 

The selection of the monomers to incorporate in the capsid should consider other items along with antigenicity, such as product release by the cells [ 15,24]. Product secretion can have a large impact on overall particle production since baculovirus-infected insect cells often exhibit proteolytic activity, which is mainly intracellular at early times post-infection [25]. The appearance of proteases often coincides with plO and polyhedrin-driven late protein production [26]. 

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