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gel shrinkage ???

agarose gel electrophoresis

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2 replies to this topic

#1 uday

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Posted 07 August 2013 - 06:45 PM

I am running some long agarose gels to be used for subsequent blotting for a Southern hybridization.  So far it was so good and suddenly I found my gel start shrinking from the top after running for about 6 hours at 40-60V. What could be the reason?

I am using 0.8% agarose gels to run restriction enzyme digested genomic DNA with 1% TAE buffer. Each time I am using new buffer. I tried with different gel boxes as well as different power supplies. I checked the buffer level too and it was well adequate to cover the gel entirely (that means no gel dehydration) but I couldnot find where I did wrong???? 



#2 hobglobin

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Posted 08 August 2013 - 07:35 AM

No real idea about this but perhaps osmotic effects when the buffer deteriorates over time or hydrated postive ions associated with the fixed anionic groups of the agarose gel migrate over time removing the hydration shell of the gel (i.e. electroendosmosis effects). Not sure if it's true.
You might try out to mix/stir the buffer or replace it after some time, perhaps this helps. Or use a buffer with more capacity such as TBE or Lithium borate.


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#3 mdfenko

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Posted 08 August 2013 - 01:03 PM

do you mean that the gell is becoming trapezoidal or shorter?

 

if shorter, are you using low-melt agarose? is the gel getting hot?

 

if trapezoidal, what is your buffer formulation? something may be different from what is normally used.

 

have you run these gels before, without shrinkage?

 

how do the bands look?


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