2 research outputs found
Genetic modification of a baculovirus vector for increased expression in insect cells
Generating large amounts of recombinant
protein in transgenic animals is often challenging
and has a number of drawbacks compared to cell
culture systems. The baculovirus expression vector
system (BEVS) uses virus-infected insect cells to
produce recombinant proteins to high levels, and
these are usually processed in a similar way to the
native protein. Interestingly, since the development
of the BEVS, the virus most often used (Autographa
californica multi-nucleopolyhedovirus; AcMNPV)
has been little altered genetically from its wild-type
parental virus. In this study, we modified the
AcMNPV genome in an attempt to improve recombinant
protein yield, by deleting genes that are nonessential
in cell culture. We deleted the p26, p10 and
p74 genes from the virus genome, replacing them
with an antibiotic selection cassette, allowing us to
isolate recombinants. We screened and identified
recombinant viruses by restriction enzyme analysis,
PCR and Western blot. Cell viability analysis showed
that the deletions did not improve the viability of
infected cells, compared to non-deletion viruses.
However, expression studies showed that recombinant
protein levels for the deletion viruses were significantly
higher than the expression levels of nondeletion
viruses. These results confirm that there is
still great potential for improving the BEVS, further
increasing recombinant protein expression yields and
stability in insect cells