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2,338 bytes added ,  09:21, 3 September 2010
Add "freeze & squeeze."
'''The following information is for basic lab technique.'''
*Also see [[DIYbio:Notebook/Safety_Manual_1.0 | the DIYbio Safety Manual]].
 
== Quick and dirty ways to do gel extraction of DNA ==
"It's called the "freeze & squeeze." When I was in grad school, this
rapidly replaced the Qiagen kits in our whole department for routine
gel purification. It works pretty well, provided a somewhat reduced
yield doesn't bother you. As it's so easy to make more DNA, it's
usually more cost-effective to use more DNA and use this MUCH cheaper
protocol. IIRC, there was an article (I'm pretty sure it was an
article and not a paper) about it in Nature or Science.
There are a lot of ways to do it, and a Google of "freeze and squeeze"
will give you a lot of results. The way we used to do it by the
following protocol:
Materials:
0.5mL centrifuge tube
1.5ml centrifuge tube
Synthetic pillow stuffing (NO COTTON, that will reduce your yield
dramatically). We used the Wal-Mart store brand stuff.
Aluminum foil
Prep:
Take a 0.5mL eppendorf tube and pierce the bottom with an 18ga
needle. Obviously, you should be very, very, very careful doing this.
Slip the smaller tube inside the larger tube and put some pillow
stuffing into the smaller tube. Put in enough to loosely fill the
bottom quarter of the tube. This is your spin column.
Method:
1) Run a gel, cut the desired band out.
2) Put it on a piece of foil and set it in the freezer (-20 works just
fine, I never noticed any improvement with lN2 or -80). Freeze it
solid, takes about an hour or so, depending on your freezer..
3) Put the gel slice on top of the pillow stuffing (no need to thaw)
4) Quick spin the elute out. Should only take 30 seconds or so; don't
overspin it.
5) If desired, clean it up with a EtOH precip or dialysis disks over
buffer.
I always had great results with this technique, and it's a hell of a
lot cheaper than Qiagen kits. Less work, too.
You can experiment with sterilizing the stuffing if you want. One guy
tried sterilizing it using a UV crosslinker, but I don't recall him
getting significantly better yields than people that just precipitated
it after the spin."
 
:: -- sgt york, Aug 24 2010, DIYbio google group
 
 

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