DNase I (RNase-free) is an endonuclease that nonspecifically cleaves DNA to release di-, tri- and oligonucleotide products with 5´-phosphorylated and 3´-hydroxylated ends. DNase I acts on single- and double-stranded DNA, chromatin and RNA:DNA hybrids.
Feature
Recombinant enzyme
Purified from non-animal host with a lower level of intrinsic RNases.
Applications
Degradation of DNA template in transcription reactions
Removal of contaminating genomic DNA from RNA samples
DNase I footprinting
Nick Translation
Quality Control
The absence of ribonucleases is confirmed by appropriate quality test. Functionally tested for digestion of template DNA after
in vitro transcription
Source: A
E. Coli strain that carries an MBP fusion clone of Bovine Pancreatic DNase I.
Molecular Weight: 29 kDa monomer
Definition of Activity Unit
One unit of the enzyme completely degrades 1 μg of plasmid DNA in 10 min at 37
oC. Enzyme activity is assayed in the following mixture: 40 mM Tris-HCl (pH 8.0), 10 mM MgSO4, 1 mM CaCl2, 1 μg of pUC19 DNA.
Inhibition and Inactivation
Inhibitors: metal chelators, transition metals (e.g., Zn) in millimolar concentrations, SDS (even at concentrations less than 0.1%), reducing agents (DTT and beta-mercaptoethanol), ionic strength above 50-100 mM.
Inactivated by heating at 65
oC for 10 min in the presence of EGTA or EDTA (use at least 1 mol of EGTA/EDTA per 1 mol of Mn
2+/Mg
2+ ).
Note: DNase I is sensitive to physical denaturation. Mix gently by inverting the tube. Do not Vortex.
