cells up overnight in 50ml culture
morning of second day check OD of cells. The OD600 should be about
1.0. Remove 10ml as time 0 and
added to 50 ml conical tube containing 100 microliter of 100mM sodium
azide (NaN3). This is to instantly kill the cells.
cycloheximide to remaining 40 mls to a final concentration of 100mg/ml.
10mg/ml stock in water (100x)
of NaN3 (final concentration 1mM) to each conical tube before
stock at 100X (100mM).
molecular weight at 65.01
10 ml of cells at specified time points. Centrifuge. Remove supernatant
and shock freeze. Store at –800C
Good starting time course:
30’, 100’, and 300’ minutes