ALKALINE MAXI-PREP FOR Ti 50, Ti 65, OR TV 865

grow 200 mls to late log
cent 5 kg x 5'
resuspend in 6 mls lysozyme sol'n (see notes for TL 100 plasmid prep)
ice x 30'
add 14 mls of .2N NaOH, 1% SDS
ice x 30'
add 14 mls of 2M NaOAc (4.8)
ice x 30'
cent 40 kg x 30'
decant supernatant, add 80 mls of 95% EtOH
ppt at -20 C
5 kg x 5', drain off EtOH completely
for Ti 50 or Ti 65
resuspend 8.7 mls ddH2O
add 8.3 g CsCl
add .9 ml EtBr (10 mg/ml)
cent 45 krpm x 36 hrs, 20 C
for TV 865
resuspend in 2 mls ddH2O
add 3.35 g CsCl
add 100 ul EtBr (10 mg/ml)
adjust final volume to 4.45 ml with ddH2O
58 krpm x 6 hrs minimum, 20 C
rebanding with CsCl
use 4.45 M CsCl
50 mM Tris-Cl (8.0)
25 mM EDTA
200 ug/ml EtBr