Smeared Western Samples

John s330477 at student.uq.edu.au
Sun Nov 11 12:08:33 EST 2001


I have had similar problems running lysates when they are viscous due to
excess DNA.
Try breaking up the DNA by passing your lysate through a small needle
syringe several times or use DNase to degrade the DNA before loading.

Hope it works!
Cheers,
John.

"Sophie" <sweiss at icr.ac.uk> wrote in message
news:20011109105429.9175.qmail at ww02.jatek.com...
> Hello,
>
> I am trying to run western blots to determine the level of protein
> expression in salmonella. I lyse the bacteria with 1% triton, 10% glycerol
> and 15 aprotonin. My protein is 46000Da and i run it on a 12% SDS PAGE
gel.
> The markers are always nicely separated and transfer to the membrane.
>
> My problem is that all my samples end up as horrible smears down the gel
> instead of compact bands. I thought it was a problem with the bacteria but
> the same thing happens when i use a highly purified version of my protein.
> Has anyone got any suggestions for why this is happening and how i can fix
it.
>
> thanks,
>
> Sophie.
>
>
>
> <http://www.biowww.net/forum/read.php?f=6&i=817&t=817>
>





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