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Star Republic: Guide for Biologists

Plaque purification of virus stock

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1. To pick a single virus from a mixture of viruses, do a Plaque Assay as has been described.

2. Mark several plaques that are well-separated from others. For generating recombinant virus, don't pick blue plaques if cells are also stained with X-gal. Some baculovirus vectors express the X-gal protein and X-gal expression will be disrupted by inserted DNA sequences.

3. Add 0.5 ml medium to a sterile eppendorf tube.

4. Pick the marked plaques by pushing the tip of a sterile Pasteur pipette through the overlay agarose. Suck the agarose into the Pasteur pipette.

5. Transfer the agarose plug to the medium by pipetting up and down several times.

6. Vortex and store overnight at 4 C to release the virus.

7. Use 100 ml to inoculate the passage one virus stock. Store the rest at 4 C.