HELPER PHAGE PREPARATION
HELPER PHAGE PREPARATION
1. Grow an overnight of NM522 in NZCYM medium.
2. Dilute overnight 1:100 and grow to an A600 = 0.3 (@2.5 x 108 cells/ml).
3. Infect cells with VCS helper phage to a m.o.i. = 0.1 (phage:bacteria = 1:10). There should be a high-titer stock of VCS in the lab.
e.g., 40ml NM522 (@1010 cells) + 10ml VCS (@109phage from a stock = 1011 phage/ml)
4. Incubate/shake @ 37oC for 8h.
5. Centrifuge 10', 15krpm to remove bacterial cells.
6. Harvest supernatant and add 0.4% chloroform.
7. Store in aliquots @ 4oC. It is a good idea to titer the prep (expect >109 phage/ml).
acc 12/90
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