1. Resuspend pellets in 100 ul of Solution I (Tris-EDTA-glucose, 50mM glucose, 10mM EDTA,
25mM Tris pH 8.0 - storage at 4?C) place on ice for 5 minutes. Add 150ul of ice-cold Solution III (for 100ml, 60ml of 5M KOAc, 11.5ml glacial Acetic Acid,
28.5ml H2O) and mix by inverting the tube gently.Grow overnight cultures of E. coli (1.5-2.0 ml) in LB broth at 37?C using appropriate antibiotic
selection.Harvest cells in microfuge tube by centrifuging for 5 minutes at 4,000 rpm, Discard media.Dry pellets (5 minutes in speedvac) and resuspend in 20-40ul TE + RNase A. Shaking for ~ 10
minutes was done to resuspend DNA pellets.Add 200 ul of freshly prepared Solution II (0.2N NaOH, 1%SDS).The solution mixture should be turbid, incubate
on ice for 5 min.Chloroform will
remove soluble proteins from the preps.2.3.4.5.6.7.8.9.10.11.12.13.