Question

agarose gel electrophoresis for DNA...
DNA in all the wells move to the same distance...why???(we used bromophenol blue as the dye and sucrose as loader in loading buffer in our lab , we ran the gel only for half of the template). and why do we use the 260/280nm ratio for checking the purity of dna.....why purity has to be checked???

Answer 1

I can at least answer to your last question : purity has to be checked because if another substance is analysed with the DNA, the eletrophoresis wil give a wrong answer. We only need to have the DNA results and not the DNA+something results. As the electrophoresis uses the electronegativity of the sample to separate them, to addition of something else will get you the wrong results if the sample is not realluy pure. It may makes the DNA go too far.

After a bit of refexion, I can say that DNA move to the same distance because, as said earlier, the technique of electrophoresis uses the electronegativity of the sample (here the DNA). If you use the same kind of substance in all the wells, they will all move to the same distance.

Answer 2

oh....actually our DNA sample was contaminated with some protein and was not completely dissolved in the buffer(it was not sufficient enough in our buffer) ...and as a standard we used lambda marker....but our isolated dna as i said earlier was with some proteins....both were upto the same mark in the template...but our dna was somewhat smeared....but to the same mark...then why the purity has to be checked??

Answer 3

Oh, I see ! Before I answer anything, I just wanna check if I understood well (I'm French, sorry, I have to be sure) : all your samples were contaminated the same way ? As all was equal, you wonder why purity has to be checked as nothing was different from one sample to one other ?