Non-DNase related smear of digested DNA? - (Aug/29/2007 )
Hi all!
Every now and then, and lately quite often, when I do a restriction digest, I just end up with a smear. The first time or two I just assumed I had been a cluts contaminated my stuff with DNases. However, the smears do not seem to be connected with a certain enzyme or buffer, it will turn up here and there with no consistent pattern. I have changed gloves, buffers, water, enzymes, tips, different DNA preps etc etc. Others, but not everyone, in the lab have seen the same thing. In my old lab, I never had anything like this happening even though I almost never used gloves and definitely never filter tips. Now, because of all this crap, I am extraordinarily anal with my digestions, but it keeps popping up anyway. 
After extensive testing, the one thing that seem to be the only common denominator are the eppi tubes! Have anyone seen anything like this before? Is there anything else but DNases that can destroy your DNA? Please help!!!
Thank you!
-Sick, tired and desperate!
is your DNAprep fresh and clean ?
any chance of star activity ?
I used to have such a problem too. How long is your digestion?
any chance of star activity ?
Have used bright new DNA preps, old ones that used to work and everything in between. I usually cut for 1.5 hrs, sometimes with enzymes that should not exhibit star activity... so I don't think that's it...
usually 1.5 hrs