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Marker migrates slower depending where is loaded


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#1 celvas

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Posted 25 August 2009 - 03:43 PM

Hi all,

today, I loaded my PCR samples, including marker ladder in first and last gel wells.
Surprisingly, the ladder migration in the last well was slower, like 100 to 200 bp less, than in the first well.
So what are really sizes of my amplification products? What is happening?

Thanks

Edited by celvas, 25 August 2009 - 08:06 PM.


#2 ivanbio

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Posted 26 August 2009 - 07:41 AM

Hi Celvas,

What you are experiencing is likely a classic physical chemistry effect = the edge that was slower likely received less current (Amperage) than the rest of the gel. These edge effects are not unusual, which is why many people recommend running the size markers in the middle of the gel (or at least right next to the sample of interest). There is little you can do about this short of running your gel very slowly and making sure there is plenty of gel between the last well and the border of your gel (and also making sure that there is an equivalent sample composition/buffer in every well because any empty well will also cause a different in current flow).

Ivan
Carlsbad, CA

#3 celvas

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Posted 26 August 2009 - 06:06 PM

Wonderful reply, thanks a lot




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