DNase I recombinant, RNase-free

Recombinant DNase I is a DNA-specific endonuclease.The enzyme catalyzes the degradation of both double- and single-stranded DNA randomly by hydrolyzing phosphodiester linkages to DNA, resulting in a mixture of oligo- and mononucleotides. All material used during the production process of DNase I recombinant is non-animal sourced, resulting in an animal-free product.

Contents

• Recombinant DNase I, RNase-free, 10 U/μl
• Incubation Buffer, 10x concentrated

DNase I recombinant, RNase-free may be used to degrade DNA in applications that are sensitive to the presence of RNase. For example, DNase I is frequently used to:

• Remove genomic DNA from RNA preparations prior to RT-PCR
• Isolate DNA-free RNA after in vitro transcription reactions
• Perform nick translations
• Map DNase-sensitive regions in eukaryotic DNA

Heat inactivation: One unit DNase I recombinant, RNase-free is heat-inactivated by 10 minutes incubation at 75 °C.

Important Note: Alternatively, DNase I recombinant, RNase-free can be inactivated and removed by phenol extraction according to standard protocols, e.g., Current Protocols in Molecular Biology.

Glycosylated form
Recombinant DNase I is heterogeneously N-glycosylated, so it appears as two bands in gel electrophoresis.
Divalent ion requirement
DNase I requires divalent cations for maximum activity. The DNA-specific endonuclease is activated by ions such as magnesium ions and is stimulated by calcium ions. Therefore, the enzyme is inhibited by metal chelating agents like EDTA.

1 kit containing 2 components

Activator: Bivalent metal ions
Working solution: Storage Buffer: 20 mM Tris-HCl, 50 mM NaCl, 2 mM CaCl₂, 2 mM MgCl₂, 1 mM dithioerythritol, 0.1 mg/ml Pefabloc SC, 50% glycerol (v/v), pH 7.6 (at 4 °C).
Incubation Buffer (10x): 400 mM Tris-HCl, 100 mM NaCl, 60 mM MgCl_2^, 10 mM CaCl₂, pH 7.9.
Enzyme Dilution Buffer: 25 mM Tris-HCl, 50% glycerol (v/v), pH 7.6 (at 4 °C).

Store undiluted enzyme solution at -15 to -25°C; storage buffer at 4 °C.

Absence of contaminants: Each lot is tested to ensure the absence of RNases and proteases according to the current Quality Control procedures.

For life science research only. Not for use in diagnostic procedures.

One unit is the enzyme activity that effects an absorbance increase of 0.001/minute under assay conditions in 1 ml at 260 nm.
Assay conditions:
Volume activity is determined according to the following assay mixture. 100 μg calf thymus DNA is incubated in 2.5 ml 1x incubation buffer with 40 to 70 units DNase I recombinant, RNase-free at +25 °C. The absorbance increase is measured at 260 nm.