DNA	> DNA MODIFYING ENZYMES	S1 Nuclease Digest of ssDNA
		Alkaline Phosphatase Digestion of DNA
		BAP
		CIP

BAP

Get a printable version of this protocol (requires Adobe Acrobat) 

To prepare BAP from the ammonium sulfate suspension its sold as, spin out the precipitate for 5 min at 4°C in microfuge, resuspend in H₂O to a concentration of 0.1 units/ul, and centrifuge desalt on Sephadex G25 equilibrated in H₂O. Adjust the desalted BAP with Tris pH 8 to 0.1 M and store at 4° C stable essentially indefinitely.

To 10 ug DNA linearized plasmid or fragments dissolved in 45 ul H₂O. Add 5 ul CB 10 X BAP, preheat to 68°C and add 1 ul BAP (0.1 units/ul Worthington). Incubate at 68°C for 1/2 hr, add another aliquot of BAP (1 ul) and continue incubation for another 1/2 hr. Then add phenol and chloroform directly to 68°C reaction volume and vortex, do multiple extractions, add NaCl to 0.2 M and ethanol precipitate as usual.

CB 10 X BAP

Tris 0.5 M pH 8.0

MgCl2 20 mM

ZnCl2 1 mM