Phage isolation

[Karl Fischer]

In article <4ncapv$3i at groa.uct.ac.za>, molapo at molbiol.uct.ac.za wrote:

Hello

For concentrating the phage, you can try the following

20% polyethylene glycol 8000
2.5M NaCl

Add 1/4th volume to your clarified culture medium, vortex, incubate at
room temperature for >1 hour, centrifuge at 4deg C at > 10,000 rpm, 30
minutes.

For DNA isolation, I would resuspend the pellet in TSE (50 mM Tris-HCl, pH
8, 150 mM NaCl, 10 mM EDTA), add SDS to 0.1% final concentration and
proteinase K to 100 ug/ml. Mix and incubate at 42-55 deg C overnight. Next
day, phenol/chloroform/EtOH as per usual.

Why the pro K, you might ask....to aid in "cracking" the virion AND to
remove any protein which might be covalently linked to your DNA (as is the
case with phages phi 29 and PRD1).

Cheers

Karl the hepB guy

-- 
Karl Fischer
tyr-2 at bones.biochem.ualberta.ca