Spin Columns Efficiency

Adam . via methods%40net.bio.net (by anonwums1 from gmail.com)
Thu Feb 4 23:42:35 EST 2010


The binding efficiency of spin columns depends on pH and salt concentration.
Generally the binding buffer you dissolve the gel slice in should have
sufficient salt and pH. The columns work really badly to bind DNA if the pH
is greater than 7.5. Some protocols suggest that you spike in a little 3 M
sodium acetate pH 5 before running it on the column. Elution off of the
column is also pH and salt dependent. You want a higher pH buffer to elute
in.

There are also binding issues with the size of the DNA fragment. There is an
optimal range of size over which the columns are supposed to work.

Generally, most of this stuff is found in the back of the manual for your
spin column. You might get some insight by calling up technical support. In
my experience, even though the manufacturer claims you get 99% recovery, you
rarely do and are lucky if you get 50-70%.

Adam

On Thu, Feb 4, 2010 at 7:41 PM, Adroit <cognizann from googlemail.com> wrote:

> Dear Bionetters,
>
>
>
> Many times I observed that before gel purification the intensity of the DNA
> band on the gel would be very high. When this intense band is sliced and
> purified by gel extraction using the spin columns its intensity decreases.
>
>
>
>
> Could any one suggest the procedure to increase the efficiency of the spin
> columns?
>
>
>
> Thanking you in advance.
>
>
>
> Regards,
>
> Adroit
>
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