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Agarose gel bands gone (for gel extraction)

DNA agarose gel bands TAE gel extraction

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#1 relevant_username

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Posted 01 May 2013 - 01:16 PM

Hi guys,

I'm really confused and sad right now. For gel extraction, I made a 0.6% agarose DNA gel, using 1X TAE + 1mM guanosine as the running buffer. I made the gel thick so that it could hold 30 uL of DNA (which I cut with restriction digest). After running the gel for some time, I checked it under UV, and I can't see anything. No marker lane. No DNA. Absolutely no trace of anything anywhere. Just nothing. It's exactly as if I didn't put anything into it, except my gel definitely has sample in it, plus a marker lane, so I'm starting to suspect aliens.

Anyone else have a similar weird experience as mine and know what caused it?

Much appreciated.

#2 relevant_username

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Posted 01 May 2013 - 01:29 PM

Okay so I did some research and basically found out that guanosine causes the DNA to be UV opaque, thus making it impossible to see in the gel! Anyone know how to remove the guanosine so that I can extract my DNA?

#3 bob1

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Posted 01 May 2013 - 02:11 PM

You could just try soaking in several changes of 1x TAE - i.e. diffusion. I don't know the limits of how much guanosine would need to be removed though, and diffusion would probably affect the band appearance before you could remove useful amounts. I think you will probably be better off re-running the gel.

I am guessing you used guanosine as a denaturant, but it isn't standard practice to run DNA gels with it in...

You could try staining with a visible light dye such as crystal violet, but I don't know that it would be compatible with gel extraction.





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