Phenol extraction of proteins
To 100 µl of PBCV-1 in a 1.5 conical plastic centrifuge tube add 150 µl of phenol.
Add DTT (~10 µl), 10% SDS (~20 µl), 10M LiCl (~15 µl) and Ponceau S (~10 µl).
Heat 5 min at 75°C, mix thoroughly.
Allow to cool, centrifuge 1 min (5K).
Draw off lower (clear) phase into 1.5 conical plastic centrifuge tube.
Add 1 ml acetone and then 5 µl of 1% Ficoll, mix.
Store 20 min at -20°C.
Centrifuge 1 min (6K).
Discard supernatant, drain tube, wash ppt once with acetone (ppt sticks to tube) and dry.
Dissolve pellet in ~50 µl cracking buffer (1% SDS; 5% glycerol; 0.03125 M Tris-HCl, pH 6.8; bromophenol blue-trace).
