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Preparing lysin

PBCV purification

Summary of lysin activities

Preparation of NC64A ghosts

Preparation of NC64A ghosts

MATERIALS:

1) NC64A Chlorella, pelleted (fresh cells preferred)
2) 70% methanol
3) Centrifuge tubes, Corex or polypropylene
4) 50 mM Tris-HCl, pH 7.8

PROCEDURE:

1) Centrifuge the NC64A Chlorella in the Sorvall centrifuge at 5,000 rpm, 5 min, 40C if fresh cells are used. If frozen cells are used, thaw the pellets and suspend with 50 mM Tris-HCl, pH 7.8. Centrifuge the cells in the Sorvall SS34 rotor or HB-4 rotor at 5,000 rpm, 5 min, 40C.
2) Decant the supernatants. Add 10-15 mL of 70% methanol to the Chlorella pellets and resuspend the cells completely.
3) Centrifuge the cells in the Sorvall SS34 rotor or HB-4 rotor at 5,000 rpm, 5 min, 40C. Decant the supernatants.
4) Repeat the 70% methanol washes 5-6 times or until the desired amount of pigment has been removed from the cells.
5) Store the Chlorella "ghosts" at 40C in 70% methanol.