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Gel Extraction Column [Orange Stain during first spin]

gel extraction cloning purification dna

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#1 jldowler

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Posted 05 September 2014 - 02:14 PM

I am completely lost on this issue.  I have used the gel extraction kit hundreds of times and never seen this.  I took my pcr product and ran it out on a 1% agarose gel.  I then used the Qiagen kit.  After adding buffer QG and dissolving my gel slices the mixture turned a deeper yellow (almost orange) color.  When I added isopropanol and spun the column i noticed that the column had stained orange.  When I ran my gel extraction product on a gel I got no band, and also my elution did not seem to freeze at -20 overnight.

 

I have tried passing buffer QG through my sample several times and washing with buffer PE several times as well.  I also have added sodium acetate as suggested by the protocol.

 

Has anyone experienced something like this before?

 



#2 bob1

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Posted 06 September 2014 - 11:01 AM

Haven't had a problem like this - but if it is a one-off, what's the problem... it may have been a dud column, so throw it out and use a new one.



#3 jldowler

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Posted 06 September 2014 - 11:46 AM

I bought a new kit and am getting the same problem.  Not sure what to make of it.   Used two different agarose supplies as well.  Maybe something in how i'm preparing my gel.







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