help w/ fluorescin GENE IMAGES kit

Arnoud van Vliet arnoud at vanvliet.u-net.com
Wed Mar 11 17:10:22 EST 1998


Hi Paul,

the last two years I worked with the Gene Images kit, and for
bacterial genomic Southerns it works fine. I can't say anything about
single-copy gene detection, but I get a strong signal after 15-50 sec
of exposure.

I might have misunderstood your questions, but it is based on
incorporation of FITC labelled nucleotides using random primer
labelling and Klenow polymerase. I gel purify my fragment, and label
>200 ng for >3h at 37 degrees C in a 50 microliter volume. Of that I
use one-third in a hybridisation.

For the antibody detection we actually dilute the antibody 1:20000,
not 1:5000 as Amersham recommends, and that gives us good Southerns.

So, we're quite happy with it. No connections or special rates from
Amersham (unfortunately)

Hope this helps,
Arnoud
arnoud at vanvliet.u-net.com

On 11 Mar 1998 19:47:28 GMT, pw at cubsps.bio.columbia.edu (Paul
Wakenight) wrote:

>If anyone out there has experience w/ Amersham Gene Images kit, I would be
>very grateful if you could tell me how well it works. We need to detect
>single copies (1pg). Also, how stable are the probes and the AP-Ab in
>reality??
>
>Thanks so much
>
>paul
>pw at cubsps.bio.columbia.edu
>




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