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


Loading limit on Agarose Gel

Loading limit on Agarose Gel - Agarose Gel Electrophoresis Forum

Loading limit on Agarose Gel - 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 02-10-2010, 12:58 AM
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Red face Loading limit on Agarose Gel



I need to digest 15ug DNA with an restriction endonuclease enzyme of 4U/ ug DNA. My Stock DNA is 2.5 ug/ul and my stock enzyme is 5U/ul.* My buffer is 10x. My gel can only allow 64 ul of the total volume of the entire reaction.* I end up having 120ul which is twice the allowable volume.* If I force this process, I will end up having a smear instead of having a good band.* I can't even divide my DNA to digest into 2.* What should I do?
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Old 02-26-2010, 10:57 AM
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Default Re: Loading limit on Agarose Gel

I think you can concentrate the DNA in a vacuum concentrator so that the volume becomes less and you have more DNA per microlitre.
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Old 05-10-2010, 11:54 AM
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Default Re: Loading limit on Agarose Gel

It is realy simple to load excessive amounts of a sample in an agarose gel. I always load something of 200-400 ul of my digested samples when I want to linearise the vector, or to liberate a chosen fragment. The trick is:
Before you cast the gel (pour melted agarose in the gel tank), cover a few wells of your gel comb with a sealing tape, there you go you have bigger well
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