DNA recovery from agarose gels-NA45
HF.JSL at forsythe.stanford.edu
Sun Jul 24 23:30:57 EST 1994
We've been NA45 membranes users for years. A couple of
modifications might be useful:
1. We generally cut a small rectangles of the gel, turn it 90o,
then electrophorese the band onto a spot. This helps in two ways.
First, closely spaced bands can be easily resolved. Second, it
reduces the area of membrane which reduces "non-specific" losses.
2. We often add yeast tRNA (Sigma) during the elution. It seems to
improve DNA yields and also gives a visible pellet, even with
nanograms of DNA.
3. We routinely do one phenol/chloroform extraction after two
sequential elutions in high salt. Although not absolutely
necessary, bits of agarose carry-over can inhibit ligation and
I still think this is one of the simplest and cheapest methods for
recovering DNA fragments. If the gel is run with EtBr you can also
see your DNA at all times (running onto the paper, e.g.).
Joe Lipsick (hf.jsl at forsythe.stanford.edu)
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