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Bands are disappearing quickly

Bands are disappearing quickly - Agarose Gel Electrophoresis Forum

Bands are disappearing quickly - Agarose Gel Electrophoresis Forum. A forum to discuss electrophoresis staining, troubleshooting, and development of films for DNA and RNA gels.


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Old 01-18-2011, 05:28 PM
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Default Bands are disappearing quickly



I am having trouble visualizing my bands in agarose gel (0.8%, 1 uL of Ethidium Bromide). I'm running PCR products (350-500bp) and I can visualize the band immediately I put it on the UV lamp (after 45' of running), but 5 mins later I canīt see almost none of them. I also tried an ethidium bath (2uL/200mL water) for 5', 10' and 30'; bands become more fluorescent with time.
And I guess I canīt see some of them from the beginning, because of this trouble and not caused by a non presence of PCR product.
Any suggestion??
Thanks a lot!!
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Old 02-23-2011, 03:47 PM
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Default Re: Bands are disappearing quickly

5 minutes under UV is actually not that fast, why would you need your gel under there for that long? If you are extracting the bands to use them you should aim to have them out in 30 seconds maximum (separating the lanes and doing them one at a time if neccessary).
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Old 02-23-2011, 11:47 PM
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Default Re: Bands are disappearing quickly

Do you have the agarose gel in some kind of solution while waiting 5 minutes?

I agree small DNAs should not be left long in gel solution, and also the quicker the extraction the better.

Also do NOT leave DNA with UV lamp ON for more than a few seconds, you will have trouble cloning or working with it later.
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