How to denature dsDNA and prevent re-annealing?
Dr. Duncan Clark
Duncan at nospam.demon.co.uk
Mon Sep 18 16:33:40 EST 2000
In article <8q5hcq$r02$1 at nnrp1.deja.com>, mbraden23 at my-deja.com writes
>Does anyone know of a simple denaturing of DNA method and reagent
>conditions to prevent re-annealing?
Alkali - simple and works.
Biotechniques vol 18. No. 3, p376 (1995)
1. incubate DNA in 0.4M NaOH, 4mM EDTA for 8 mins at 37C
2. Neutralise with 0.1volume of 2.5M NaAcetate pH 5.2
3. Ppt by adding 3 volumes EtOH.
4. 70% ethanol wash the pellet.
Works for plasmids and seems to be OK for linear DNA.
Working the night shift!
The problem with being on the cutting edge is that you occasionally get
sliced from time to time....
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