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Dear Narayanan, you could simply test that by a negative control: transform your cells with an empty vector, and follow the same (exact) purification protocol. No DNase activity there = probably no DNase contaminant in your original prep. Cheers Filip On 8/6/07, Narayanan Ramasubbu <[log in to unmask]> wrote: > > Dear All: > This is off topic but I would rather try here first. > I am wondering whether Dnase could be a contaminant during the nickel > affinity purification of a his.tag protein expressed using pET29b. The > cells were disrupted using sonication only. Very high yield (30 mg/liter > of cell culture). FPLC purification. SDS-PAGE shows a singe band even > when overloaded. Silver staining is in progress. > The protein I am interested in does not have any similarity to Dnase. > However, when treated with supercoiled dna, the dna is degraded as > efficiently as with Dnase I from Sigma. > > I am looking for ways to show that my preparation does not have any > Dnase contamination. I would appreciate it very much if someone points > me in the right direction. > Thanks a lot > Subbu > -- Filip Van Petegem, PhD Assistant Professor The University of British Columbia Dept. of Biochemistry and Molecular Biology 2350 Health Sciences Mall - Rm 2.356 Vancouver, V6T 1Z3 phone: +1 604 827 4267 email: [log in to unmask]