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UV-Imaging Chambers: Oversaturation from many gel lanes?


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#1 djvan

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Posted 09 April 2016 - 10:40 AM

Hi All,

 

I've been running some massive DNA-agarose gels in a screening application downstream of PCR.  The gels contain 4 combs of 50 lanes each.  The center of the lanes within a comb are separated by about 5mm.  I cut the gel prior to imaging, so that only 50 lanes are imaged at a time.

 

I think I'm running into some problems with these large gels in relation to the UV signal.  I think I might not be detecting weak (low concentration) DNA bands on these gels.  Can bright DNA bands in adjacent lanes 'mask' weak DNA bands lanes?  The camera needs to be zoomed-out pretty far to encompass all of the lanes, and I think I might be losing sensitivity?  I noticed if I crank up the exposure, weaker bands do appear, but the other, stronger bands, become oversaturated as does my DNA ladder.

 

I could load more of these weaker samples, but the issue is I will not know which lanes need to have more loaded until after I run the first gel.  It would be ideal to only have to run the gel once, as this is a screening process and quite laborious.

 

Thanks for any help!



#2 phage434

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Posted 11 April 2016 - 06:41 AM

You can image each gel twice, once at low and once at high exposure.






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