DNA control on Northern?

Hiranya Roychowdhury hroychow at NMSU.EDU
Mon Nov 16 11:01:16 EST 1998

At 10:29 AM 11/16/98 +0100, Frank Maiwald wrote:
>Dear all,
>I would like to use a fragment of lambda DNA (part of a cloned insert)
>as a positive control in a Northern blot. In my first attempt this
>*positive* control did not give any signal, although the target DNA
>quantity should be more than enough. I denatured the DNA at 95 C for 10
>minutes ice shocked it and loaded it with RNA loading buffer on a
>formaldehyde/ MOPS/ agarose gel.
>Is it possible that the DNA renatures in the loading buffer or gel? Is
>is degraded? Or does my result simply mean what you think at first sight
>- the probe does not hybridize to the control?
>Any thoughts or hints are welcome; thanks in advance,

Here's a hint...
You see, the Formaldehyde in the gel keeps the RNA denatured (the heating of
RNA and the formamide in the sample buffer denatures the RNA species prior
to loading). So....

Dr. Hiranya Sankar Roychowdhury
Plant Genetic Engineering Lab.
New Mexico State University
Las Cruces, NM 88003
Ph. (505) 646-5785
hroychow at nmsu.edu

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