Removal of CsCl from CsCl-EtBr gradient preps
Conrad Halton Halling
chh9 at quads.uchicago.edu
Mon Sep 9 11:53:00 EST 1991
In article <0094E602.8B8D6680 at aclcb.purdue.edu>
muriana at aclcb.purdue.edu (murianap) writes:
>I am posting to inquire if anyone can give me a easy way of getting rid of
>the CsCl remaining in EtBr-extracted CsCl chromosomal preps? I have
>routinely used Amicon Centricons for plasmid preps, but believe these will
>get clogged if I use them for chromosomal preps. I've tried the method in
>the new Maniatis (add 3 vol d.H20 + 2 vol EtOH) but that didn't work. Is
>there any other method besides dialysis?
>Thanks in advance,
>Peter M. Muriana
> Name: murianap
>Internet: muriana at aclcb.purdue.edu
> Phone: 317-494-8284
I think the description in the "new Maniatis" (Sambrook et al., 1989.
Molecular Cloning, 2nd ed., p. 1.46) is just plain wrong. Here is what
I routinely use for plasmid preps -- it should work for chromosomal preps too:
1) Measure the volume of the DNA + CsCl solution.
2) Add two original volumes of TE or dH2O and mix.
3) Add six original volumes of 95% ethanol and mix.
The DNA will precipitate at room temperature. I spin at 12,000 x g for
15 minutes to recover the DNA.
Example: For 1 ml of DNA + CsCl solution, add 2 ml of TE and mix, then
add 6 ml of 95% ethanol and mix.
chh9 at midway.uchicago.edu
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