Flowchart-Southern blot labs (2 days)
Monday
Traditional blot (gel) / Colony lift· Restriction digests (TAs prepared)
· 5 gels (TAs prepared)
· Run digests on gel at 50mV (1 group per gel), 2 h
· Make probe (while the gel runs), let stand over night (set up 20 minutes)
· Prepare gel for blot (1.5 h)
(cut paper towels etc, during the wait) / · Colony lift and transfer of DNA onto membrane (30 minutes), auto cross-link (store membrane until Tuesday)
· Set up over-night blot (10 minutes)
Tuesday afternoon (11am-2pm)
Traditional blot (gel) / Colony lift· Dissemble over-night blot and cross link membrane
· Pre-hybridize both membranes (30 minutes- 1h), and transfer to hybridization solution, leave over night
Wednesday
Traditional blot (gel) / Colony lift· Stringency washes (45 minutes)
· Blocking and detection (2-2.5 h)