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Synaptic Proteins: Protocols

Welcome to the Home of Vesicle Trafficking

A small repository of synaptic protein information

Protocols

Caution! In converting these protocols to HTML, some units have been scrambled.
Specifically, microliters (ul) often appear as milliliters (ml). Think first!


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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