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Dotted bands on my DNA agarose electrophoresis gel.


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#1 Rje

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Posted 11 October 2013 - 06:03 AM

I have uploaded a picture of the gel I have just run. I have been attempting to create two overlapping fragments, which together encompass the entire mitochondrial genome (both fragments being between 8000 - 9000bp in length), using long range PCR. The samples on the gel are the products of 4 of these PCRs. I am confused because it the area where I would expect to see the bands which equate to my wanted fragments, I have dots. I haven’t seen this before, can anyone help explain this? Thank you. (My gel was 0.5% agarose, 7X10cm, run at 45v for 3 hours)

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#2 dimensionsbio

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Posted 12 October 2013 - 04:20 AM

I have seen rounded or dot-like bands before using EtBr when there is a lot of DNA present in the band.  The lanes on the gel appear to be overloaded as the smearing above and below your bands of interest indicate a loss of resolution.  Have you tried to extract the fragments and then run a diluted sample or smaller load on a gel?


Edited by dimensionsbio, 12 October 2013 - 04:21 AM.


#3 Rje

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Posted 13 October 2013 - 11:12 PM

Ah ok, that is probally the reason then. Yes, I think the concentration of DNA is quite high. I haven't extracted fragments and then run a diluted sample or smaller load on a gel yet, but maybe i could try this. Thank you for taking the time to reply to my question.






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