Agarose gel electrophoresis uneven migration - (Oct/20/2014 )

I  use 1.3% agarose for my restriction digestion, it comes out wavy and irregular this started happening recently what could be the reason and what should I do?

You may not be completely dissolving the agarose. Make sure you heat it completely and swirl to mix it after heating. It should be crystal clear and uniform.

There could also be a non-uniform depth of agarose in your gel, or incomplete coverage of the gel by buffer.

phage434 on Mon Oct 20 18:00:05 2014 said:

You may not be completely dissolving the agarose. Make sure you heat it completely and swirl to mix it after heating. It should be crystal clear and uniform.

There could also be a non-uniform depth of agarose in your gel, or incomplete coverage of the gel by buffer.

Thanks for the reply

I'm doing all those basic steps, the curving appear only for restriction while pcr product gel is fine.

too much salt in the slots?

hobglobin on Tue Oct 21 17:36:42 2014 said:

too much salt in the slots?

Thanks for the reply

I use promega agarose powder and pre made 10xtbe buffer that I dilute to 1x with distilled water.  

By too much salt, Hobgoblin means that there is probably too much salt in the sample, as a result of running the digest in the digestion buffer.

actually I meant the restriction mix you pipet in the slots which may have a too high salt concentration. Also the restriction enzymes still bound to DNA can result in such migration patterns.

Edit: too late...thanks bob

hobglobin on Tue Oct 21 19:39:42 2014 said:

actually I meant the restriction mix you pipet in the slots which may have a too high salt concentration. Also the restriction enzymes still bound to DNA can result in such migration patterns.

Edit: too late...thanks bob

I use  newengland biolabs cutsmart which includes buffer tube and restriction enzyme tube.

you may be seeing the effects of a slight shift in pH due to the buffer used in the reaction.