sodium acetate in sequencing gel buffers!
drm21 at mole.bio.cam.ac.uk
Thu Mar 6 05:25:01 EST 1997
In article <331E2908.4E13 at dept.agry.purdue.edu>, zarir
<zarir at dept.agry.purdue.edu> wrote:
>i want to know the reason for adding sodium acetate to the lower buffer
>tank while doing manual sequencing. I know it reduces compressions, by
>altering the voltage gradient, but how? what is the chemistry involved?
>Please reply urgently.also some reference where it is documented.
>dept. of agronomy,
>zarir at dept.agry.purdue.edu
AFAIK, sodium acetate has absolutely no effect on compressions. What it
does is to make bands towards the bottom of the gel run more closely
together (as they do in gradient gels). By making the bands at the bottom
run closer, you can run the gel longer before the first bands run off, and
read lots more bases.
You are right about the voltage gradient - by increasing the ionic strength
at the bottom of the gel, you decrease the resistance (R). Since V=IR, and
I is constant(*) in a series circuit (which this is), the voltage drop (V)
across the bottom part of the gel decreases and the bands slow down. ie The
V/cm across the top part of the gel is higher than the V/cm across lower
Sorry, I haven't got a reference, but you could look in the archives (and
possibly the FAQ?) as its been discussed here fairly extensively.
(*) Actually of course, if you run the gel at constant power, the overall
current (I) will change with time - but at each instant it will be the same
through each cm of the gel....
D.R.Micklem, Time flies like an arrow...
Wellcome/CRC Institute, Fruit flies like a banana.
Cambridge CB2 1QR, UK Tel: [+44] (0)1223 334129
Email:drm21 at mole.bio.cam.ac.uk Fax: [+44] (0)1223 334089
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