Jul 7 2010

How flashBAC works

 The flashBAC system works by creating a recombinant baculovirus to overexpress the protein encoded by your ‘gene of interest’ in a cell line of your choice. The flashBAC TEN, flashBAC GOLD and flashBAC ULTRA products also contain gene deletions to help further enhance recombinant protein yield and quality.

A recombinant baculovirus is produced by simply co-transfecting insect cells with the flashBAC virus DNA supplied in the kit and a suitable transfer vector (such as a pOET plasmid) containing ‘the gene under investigation’. Homologous recombination within the insect cells (see diagram above) restores the function of an essential gene that is partly deleted in flashBAC (ORF1629), allowing the flashBAC virus DNA to replicate and produce virus particles. This also simultaneously inserts ‘the gene under investigation’ into the virus DNA under the control of the polyhedrin promoter. The recombinant virus genome, with the restored essential gene, replicates to produce baculovirus that can be harvested from the culture medium of the transfected insect cells (and forms a seed stock of recombinant virus). This recombinant virus is used to produce your protein.

As it is not possible for non-recombinant virus to replicate there is no need for any selection system. Additionally we also supply a control vector with each kit so that you can double check that your co-transfection has worked. This one-step procedure greatly facilitates the high throughput production of baculovirus expression vectors via automated systems. However, it is also of benefit to the small research group just requiring one or a few recombinant baculoviruses prepared in individual dishes of cells.

 To get started with the flashBAC system all you need is a flashBAC kit and a transfer plasmid into which you clone/have cloned your gene of interest. We offer our own range of pOET transfer plasmids and the flashBAC system is also back compatible with all baculovirus transfer vectors based on homologous recombination in insect cells at the polyhedrin gene locus. Or follow the link to view our expression vectors page.

 

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Fax: +44(0)1865 483250
Email: info@oetltd.com
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