Lysin prep - SDS gel
Three 100ul aliquots of lysin prep (dialyzed overnight against 0.1M ammonium acetate) were placed in 1.5ml microfuge tubes. The first was untreated. To the second was add 1ul 5mg/ml CTAB and to the third 2ul 5mg/nl CTAB. Samples were incubated 15 min at 37C and centrifuged 3 min at 14,000 RPM. Supernatants (no pellets were actually observed) were diluted to 0.5ml with DW and precipitated with TCA. Precipitates were taken up in 30ul of pH 9 borate cracking buffer with 1mM iodoacetamide and then heated 2min at 100C. Aliquots were applied to gels along with aliquots of purified virus treated similarly with cracking buffer.
Clearly CTAB doesn't interfere with TCA ppt. The improved yield with CTAB could reflect either facilitated extraction of lysin from DNA (note tops of gel lanes) or preventing the lysin DNA complex from pelleting at 14,000g (despite no obvious pellet).
