DNA stuck in wells

Paul N Hengen pnh at fcsparc6.ncifcrf.gov
Mon Jan 9 16:09:05 EST 1995


In article <3enkg7$68i at dns1.NMSU.Edu> smori at nmsu.edu (Shahram Mori) writes:

>: I've tried the 4% non-deturing gel (a band-shift assay) with TBE and TAE,
>: with Scholler buffer and BCA and the probe (144bp) doesn't seem to move
>: any distance into the gel even in the lane with just buffer and probe.
>
> Do you heat up your DNA prior to loading?
> It maybe that 2ndary structures might prevent it from entering. I boil for
> 2min and place on ice immediately -> load.

I don't recommend this because it will certainly kill the gel-shift experiment.
The shift assay is based on protein binding to DNA causing a change in mobility.
Since the DNA in buffer looks the same, I suggest changing buffer conditions.
The DNA in buffer should give a nice tight band on the gel.

P.S. What is "Scholler buffer" anyway?

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