DNase I (EC: 3.1.21.1), is a recombinant endonuclease that cleaves both single- and double-stranded DNA preferentially at phosphodiester linkages adjacent to pyrimidine nucleotides, producing oligodeoxyribonucleotides with 5'-phosphate and 3'-OH ends. The catalytic activity of DNase I is dependent on metal ions including structural calcium Ca2+ and catalytic magnesium Mg2+ and manganese Mn2+. In the presence of Mg2+, DNase I cleaves each strand of dsDNA independently and randomly. In the presence of Mn2+, DNase I cleaves each strand of dsDNA at approximately the same sites.
Applications:
Technical notes:
Applications:
- It is typically used for selectively degrading unwanted DNA in the presence of RNA in the samples.
- DNase I foot printing assay for identification of the precise DNA sequence bound by the protein of interest
- Removing genomic DNA from RNA samples prior to sensitive applications such as RT-PCR (Reverse Transcriptase - Polymerase Chain Reaction), and Northern blotting
- Removing DNA template after in vitro transcription
- Radioactive or fluorescent labeling: digesting the radioactively labeled or fluorescently labeled DNA fragment, and then uses gel electrophoresis to detect the resulting cleavage pattern.
- DNA removal in bioprocessing applications, like DNA removal from cell lysates to decrease viscosity providing better yields
- Applying to experiments like Nick Translation, Actin binding, UV crosslinking of proteins to nucleic acids, and investigation on chromatin
- Digesting DNA in the tissue dissociation medium from cell damage.
Technical notes:
- Purity >98% by Coomassie Blue Staining
- Specific activity 5000/1000 U/mg or 5000/1000 U U/ml
- Endotoxin ≤ 0.1 EU/ml (0.0001 EU/ 1000 U)
- Unit Definition: One unit is defined as completely degrades 1 μg of DNA substrate in 10 min at 37 oC in the reaction buffer.
- Reaction buffer: 50 mM Tris-HCl in at pH 8 with 2 mM MgCl2 and 5 mM CaCl2.
- Storage and Shipping Stored in the buffer of 50 mM Tris-HCl in 50% glycerol at pH 8 with 2 mM MgCl2 and 5 mM CaCl2.