04716728001

DNase I recombinant, RNase-free

from bovine pancreas, expressed in Pichia pastoris

• Enzyme Commission number: 3.1.21.1 (BRENDA, IUBMB)

Properties

• biological source: bovine pancreas
• Quality Level: 100
• recombinant: expressed in Pichia pastoris
• form: solution
• mol wt: ~39 kDa
• packaging: pkg of 10,000 U
• manufacturer/tradename: Roche
• optimum pH: 7.0-8.0

Description

General description

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

Specificity

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.

Application

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

Features and Benefits

• Eliminates DNA contamination from any RNA sample
• Contains no detectable RNase or protease activity
• Can be heat inactivated, thereby eliminating the need for organic extraction
• Shipped with an optimized incubation buffer, which supports maximum DNase activity
• Produced via an entirely animal-free process, to eliminate any risks associated with animal-derived material

Packaging

1 kit containing 2 components

Quality

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

Specifications

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.

Unit Definition

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.

Preparation Note

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).

Storage and Stability

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

Other Notes

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

Safety Information

• Storage Class Code: 12 - Non Combustible Liquids
• WGK: WGK 1
• Flash Point(F): does not flash
• Flash Point(C): does not flash