DNA elution from gels
Ross.Thomas at med.monash.edu.au
Mon Feb 20 19:46:36 EST 1995
pnh at fcs260c2.ncifcrf.gov (Paul N Hengen) wrote:
> In article <3gbs9n$n6t at rebecca.albany.edu> labonnes at csc.albany.edu
> (S. LaBonne) writes:
> |: We're trying to elute individual bands from gels of PCR products for use
> |: in sequencing. We've been using electroelution, but wonder what is the
> |: fastest and/or most reliable way to get DNA out of a gel band for this
> |: purpose?
> | I assume you mean agarose gels. Having tried lots of methods for
> | extracting bands from agarose gels, my current favorite is digestion
> | with GELase (Epicentre Technologies, Madison, WI; 1-800-284-8474).
> | In my hands it reliably gives very good yields of clean DNA, good for
> | sequencing, cloning or whatever.
> Have you been able to PCR amplify DNA extracted in this way? Past discussions
> here concluded Gelase extracted DNA does not work for PCR. I assume you are not
> cycle sequencing :-o
My favorite method is to simply freeze the gel slice in an eppy using
liquid N2, spin for 5 min and take the super, then precipitate in ethanol
and salt (Sodium acetate for example).
You will lose about 50% of the DNA this way (depending on the fragment
size), but the DNA is clean and intact. This is great for ligations
and I expect for PCR also, although I haven't tried. This method is
gentle and you don't add anything that you must get rid of later.
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