Monobac System–A Single Baculovirus for the Production of rAAV

Large-scale manufacturing of rAAV is a bottleneck for the development of genetic disease treatments. The baculovirus/Sf9 cell system underpins the first rAAV treatment approved by EMA and remains one of the most advanced platforms for rAAV manufacturing. Despite early successes, rAAV is still a complex biomaterial to produce. Efficient production of the recombinant viral vector requires that AAV replicase and capsid genes be co-located with the recombinant AAV genome. Here, we present the Monobac system, a singular, modified baculovirus genome that contains all of these functions. To assess the relative yields between the dual baculovirus and Monobac systems, we prepared each system with a transgene encoding γSGC and evaluated vectors’ potency in vivo. Our results show that rAAV production using the Monobac system not only yields higher titers of rAAV vector but also a lower amount of DNA contamination from baculovirus.

Use of Bacterial Artificial Chromosomes in Baculovirus Research and Recombinant Protein Expression: Current Trends and Future Perspectives

The baculovirus expression system is one of the most successful and widely used eukaryotic protein expression methods. This short review will summarise the role of bacterial artificial chromosomes (BACS) as an enabling technology for the modification of the virus genome. For many years baculovirus genomes have been maintained in E. coli as bacterial artificial chromosomes, and foreign genes have been inserted using a transposition-based system. However, with recent advances in molecular biology techniques, particularly targeting reverse engineering of the baculovirus genome by recombineering, new frontiers in protein expression are being addressed. In particular, BACs have facilitated the propagation of disabled virus genomes that allow high throughput protein expression. Furthermore, improvement in the selection of recombinant viral genomes inserted into BACS has enabled the expression of multiprotein complexes by iterative recombineering of the baculovirus genome.