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first time running DNA polyacrylamide TBE gel


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#1 labstud4

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Posted 20 August 2014 - 04:30 PM

This is my first time running nucleic acids on a polyacrylamide TBE gel. I first cast a polyacrylamide gel using our BioRad equipment that we usually use for Westerns (1X TBE, 10% 29:1 acrylamide, 0.1% APS, TEMED), loaded my samples, and ran at 130 V for 70 minutes in 1X TBE. I noticed that my bromophenol blue marker only migrated one-third of the way down the gel. Could this be due to using loading dye I usually use for TAE agarose gels (1X TAE, Ficoll 400)? Should I make another loading dye with 1X TBE for these gels? Will my samples still migrate okay? Thanks.



#2 mdfenko

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Posted 21 August 2014 - 03:40 AM

yes, you should use loading dye with tbe. the tae may have an effect on migration.


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#3 labstud4

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Posted 21 August 2014 - 08:30 AM

If I am using BioRad containers, do I need to pre-electrophorese the gel?



#4 Trof

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Posted 21 August 2014 - 08:39 AM

The bromphenol migrates also depending on the gel density, different on 1%, 2% agarose gel. So I would expect it to migrate very differently on polyacrylamide gels.
And you shouldn't use different buffers in loading and in gel. 


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#5 mdfenko

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Posted 22 August 2014 - 03:43 AM

If I am using BioRad containers, do I need to pre-electrophorese the gel?

it's always a good idea to pre-run a continuous buffer gel like the one you're using (it's not a good idea when running discontinuous gels), it "cleans" the gel.


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