3 replies to this topic

[Raygoza]

Now, the question title may sound weird right off the bat, but my question is as follows:

 

Some restriction enzymes warn about the DNA mobility changing properties of some enzymes because the enzyme may remain attached to the DNA, so my question is can i add SDS to the loading dye and perhaps resolve this problem to detach proteins off the DNA ?  i wanted to make sure that SDS doesn't have DNA denaturing properties as well. or if the idea sounds reasonable.

 

I know that you may inactivate some of the enzymes before electrophoresis but please focus on my question as it is.

 

Thanks

Edited by Raygoza, 07 September 2015 - 04:49 AM.

[bob1]

I think for SDS to work you would need to heat the sample to about 70 C, which would probably denature (some) DNA and might result in further migration problems.

[hobglobin]

NEB suggests to add 0.5.-1 % SDS to the loading buffer to dissociate the enzyme from the DNA when you have such problems, therefore it should work.
DNA will not be denatured except you will heat it or add chemicals such as formamide or dimethyl sulfoxide. The mechanism and result of denaturation in DNA is quite different compared to proteins....and in DNA usually reversible.

[Andrea Fortina]

I saw DNA loading buffer from Thermo contains 1% SDS, which could eliminate DNA-protein interactions, prevent appearance of additional bands due to annealing of DNA molecules with cohesive ends.