Ethidium Bromide or safe dyes for DNA??? - (Oct/22/2010 )
seanspotatobusiness on Wed Nov 10 15:55:49 2010 said:
Why can't you add EtBr directly to samples before loading, supposing it's convenient (e.g. a loading dye which also contains EtBr)?
Yeah, good question. If you run EtBr pre-stained gel before, you will know the EtBr will move towards the opposite direction with your nucleic acid. So, if you mix it to your sample with loading dye, you will see nothing in the end because EtBr had moved out of the gel.
Unless somebody proves me wrong...
-adrian kohsf-
adrian kohsf on Wed Nov 10 16:41:28 2010 said:
seanspotatobusiness on Wed Nov 10 15:55:49 2010 said:
Why can't you add EtBr directly to samples before loading, supposing it's convenient (e.g. a loading dye which also contains EtBr)?
Yeah, good question. If you run EtBr pre-stained gel before, you will know the EtBr will move towards the opposite direction with your nucleic acid. So, if you mix it to your sample with loading dye, you will see nothing in the end because EtBr had moved out of the gel.
Unless somebody proves me wrong...
Well it is suggested sometimes (here) and you even can buy ready-made loading buffers containing Etbr, e.g. here, though I guess this is a waste of money...
This paper may help, though it is not online:
Angermüller SA, Sayavedra-Soto LA: Rapid visualization of genomic DNA and total RNA in agarose gels. Biotechniques. 1990 Jan;8(1): 36-38.
If someone has it as pdf, I'd like to have a copy...
-hobglobin-
adrian kohsf on Wed Nov 10 16:41:28 2010 said:
If you run EtBr pre-stained gel before, you will know the EtBr will move towards the opposite direction with your nucleic acid.
This is undoubtedly true for EtBr that is not associated with DNA, but I'm pretty sure once it's intercalated, it'd be carried along with the DNA molecule...
-HomeBrew-
The intercalated EtBr is carried along with the DNA, but it does retard migration a bit so it is not good to do if you want to compare sizes of migration accurately.
-bob1-
Wow, interesting. If this is the case, I will try to run a gel tomorrow, one without EtBr stained but add EtBr in the sample buffer; another as normal EtBr stained gel...
will let you guys know the result.
-adrian kohsf-