Etoh wash - confessions of a paranoid researcher....
nash at biologysx.lan.nrc.ca
Wed Sep 1 09:43:14 EST 1993
In article <1993Sep1.074325.19613 at gserv1.dl.ac.uk> daj at uk.ac.ic.nhm ((David Johnston) daj) writes:
>From: daj at uk.ac.ic.nhm ((David Johnston) daj)
>Subject: Re: Etoh wash - confessions of a paranoid researcher....
>Date: Wed, 1 Sep 1993 08:40:57 GMT
>>When you do an ethanol wash of a DNA precipitate do you try to resuspend the
>>pellet and spin, rinse the pellet and spin, rinse the pellet and just dry...
>You'll never truely resuspend the pellet 'cos its a precipitate, but you
>may dislodge it from the tube wall and break it into bits if it is big and
>you vortex it like hell. Also, often, even with gentle handling, 70%
>ethanol appears to dislodge pellets from the tube wall. So.....
>(1) If you can see your pellet clearly and it is still attached to the wall
>after addition of the 70% and a gentle mix/vortexing (remember, the aim of
>the rinse is to help flush out salts so mixing must help) then there is
>probably no need to spin, but if your pellet is small/invisible/loose then
>what is the "cost" of an extra minute or two in the microfuge against peace
>of mind (as the DNA is already precipitated into "lumps" (whether you can
>see them or not) you only need to do a brief respin).
I know mixing will help, but I haven't mixed in years, just added the 70%
ethanol, and then sat it on the bench for a moment, then centrifuged it for
60 sec, decanted supernatant, and used. Two different things I do.. I
aspirate off the supernatant with a drawn out Pasteur pipette end (hooked
up to a water-line vacuum). You never lose pellets that way - at worst,
they stick to the glass tip. It gets out much more of the ethanol. Also, I
do a final rinse in absolute ethanol. It seems to dehydrate the pellet, so 2-
5 min on the bench is enough to dry out the pellet before adding the TE or
water or whatever.
John Nash | Email: Nash at biologysx.lan.nrc.ca.
Institute for Biological Sciences |
National Research Council of Canada | Email to my other NRC accounts
Ottawa, Ontario, Canada. | is usually forwarded here.
*** Disclaimer: All opinions are mine, not NRC's! ***
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