Darren Tyson wrote:
>> Scott McMahan wrote:
> >
> > I was having problems recently with the samples of my Tris-Tricine
> > SDS-PAGE/Westerns streaking vertically. After checking the usual suspects
> > (old sample buffer, precipitated sample, etc.) I found a few protocol
> > books mentioning that turning the current down sometimes helps this
> > problem. When I did that, it did drastically decrease the streaking, but
> > really increased the running time. I was wondering if anyone knew what
> > phenomenon is causing this streaking, and is there some other way of
> > correcting it so I can run my gels faster again.
> >
> > --
> > Scott McMahan
> > mcmahan at oncology.wisc.edu>> Scott,
>> In my samples the cause of streaking (the majority of the time) appears
> to be excess unfragmented DNA. To fix this I sonicate and boil samples
> longer before loading on the gel.
>> Cheers,
>> Darren
> --
> Darren Tyson
> Ph.D. Candidate in Cell and Molecular Biology
> Saint Louis University
>tysondr at slu.edu or darren at primary.net>http://www.scimart.com/darren/
Why don't you add a little bit of DNAse I? For the production of enzyme
from bacterial cells, we add ~2 - max. 5ug/L of DNAseI per liter of cell
broth after or sometimes even before the cell disruption step; 20min at
0C is sufficient. At RT or higher you need even less.
This is so low, you will not see it in your gel, and the enzyme is very
cheap.
Achim
