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fainting of PCR amplification bands with time
Posted by: mtfelicio (IP Hidden, New member, 1)
Date: July 11, 2005 04:12PM
I have been doing RAPD's and running the PCR amplification products on 1.2%agarose gels at 80 mV and get nice bands after ca. 1 hour run time.
Usually I want to leave the gel running for more time (another hour) so that the fingertyping profiles are clearer and bands are more apart one from each other. But when I leave the gel running for more time unfortunately I get bands that are very faint as if they were less focused and as if they are beginning to disappear. Any possible suggestions or comments on this will be most helpful. Thanks a lot in advance Best regards Teresa
Re: fainting of PCR amplification bands with time
Posted by: Marc (IP Hidden, Unregistered user, )
Date: July 15, 2005 02:26PM
It sounds like you are running it so long that your ethidium bromide is literally leaving the gel. From what I understand, ethidium bromide is attracted to the (-) pole, and so after time it will abandon your gel. Just add a few more uL of ethidium bromide to the (stagnant) running buffer and allow some time for it to enter the gel. The resolution should improve
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