DNA Preparation - Blunt-Ended DNA Fragments


Jeff Lawrence

5' Overhangs

1. Extract and precipitate DNA. Resuspend in: 
	77 µl	ddH2O
	9 µl	10X Medium Salt Buffer
	4 µl	40 mM dNTP (10 µM each)
	1 unit	Klenow
2. Incubate at 25° for 30 min.

3. Heat to 70° for 5 min to inactivate enzyme.

4. Extract and ethanol precipitate.


3' Overhangs

1. Extract and precipitate DNA. Resuspend in: 
	17 µl	ddH2O
	1 µl	10X Medium Salt Buffer
	2 µl	20 mM dNTP (5 µM each)
	1 µl	T4 DNA Polymerase (@ 5000 unit/mL)
2. Incubate at 37° for 15 min.

3. Heat to 70° for 5 min to inactivate enzyme.

4. Extract and ethanol precipitate.
Last Update: Thursday, 12-Dec-2024 00:55:19 UTC
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Eric Kofoid eckofoid at ucdavis.edu