Large Volume Marine Cyanophage Phage Purification
Grow up cells in large volumes (1-2L) to yield ~1011 - 1012 cells using 'microbubbling' technique.
Add Rnase A (10ug ml-1 = for 2L, 250µl of Qiagen's 100mg/ml stock) and DNase1 (0.25 SU ml-1 =2L, add 50µl of stock).
Centrifuge 12,500 rpm (23,975xg) in Beckman Ja-14 for 15 minutes at 4°C Filter the supernatant through a Kim Wipe tissue paper.
Mix in polyethylene glycol (PEG 8,000; 100g L-1) by gentle stirring.
Incubate overnight at 4°C.
Collect PEG-phage precipitate by centrifuge (6 tubes at a time for the 2L volumes) at 12,500rpm (23,975xg) in Beckman Ja-14 for 15 minutes at 4°C.
Discard supernatant.
Gently resuspend pellet into ~1/150 volume Pro99 (~1-2 ml).
Add in resuspension media -place the tube on its side in syrofoam cut-outs so that it rests at an angle.
Put on orbital shaker (150 rpm) in cold room for overnight re-suspension.
Combine resuspended PEG concentrated phage into a single tube (now ready for layering onto a CsCl gradient).
Prepare a CsCl step gradient (1.3, 1.4, 1.5, 1.65 bands) in ultracentrifuge tubes.  
Add concentrated phage sample at top.
Spin 28,000 rpm for 4 hours at 4°C in SW28 swinging bucket rotor.
Remove purified phage band (interface of ρ = 1.4 and 1.5 bands on step gradient). 
Step dialyze in Slide-A-Lyzer dialysis cassettes (Pierce #66425; 10K MWCO, 0.5-3.0 ml volume) - 30 minutes against 5xNaCl (3M), 3xNaCl (1.8M), two changes of 1xNaCl buffer [MTM100 = 600 mM NaCl, 100 mM TrisHCL (pH = 7.5), 100 mM MgCl2].  
Store dialyzed particles at 4°C until needed. 
Incubate for 1 hour at room temperature. 
Add 116g NaCl per Liter of lysate (thus 2M addition + seawater yields final concentration ~2.6M).
Incubate on ice 30 minutes.
