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| Product Name | DNase I, RNase-Free Enzyme |
| Catalog No. | NATR-HMM-0123 |
| Description | A highly purified bovine pancreatic deoxyribonuclease I (DNase I) preparation that has been rigorously tested to be free of detectable ribonuclease and protease activity. The enzyme is supplied lyophilized with an optimized reconstitution and reaction buffer for the removal of contaminating DNA from RNA preparations and other applications. |
| Intended Use | Elimination of genomic DNA contamination from RNA samples prior to reverse transcription; removal of DNA templates from in vitro transcription reactions; DNase I footprinting of DNA-protein interactions; nick translation for probe labeling; and degradation of DNA in protein purification workflows. |
| Principle / Technology | DNase I is an endonuclease that non-specifically cleaves single-stranded and double-stranded DNA by hydrolyzing phosphodiester bonds, producing 5ʹ-phosphorylated oligonucleotides with 5ʹ-phosphate and 3ʹ-OH termini. The enzyme requires divalent cations (Mg2+ for single-strand nicks, Ca2+ and Mn2+ for double-strand cleavage). |
| Detection Method | Agarose or polyacrylamide gel electrophoresis with DNA staining to verify DNA removal; PCR amplification of genomic targets as a highly sensitive assay for residual DNA; spectrophotometric and fluorometric quantitation of DNA before and after treatment |
| Sample Type | Total RNA preparations from any source (tissue, cell, bacterial, plant); purified mRNA; in vitro transcription reaction mixtures; any nucleic acid sample requiring DNA removal |
| Performance Range / Specifications | Enzyme activity: ≥2,000 Kunitz units per mg protein; specific activity verified by hyperchromicity assay using calf thymus DNA at 25°C; one unit degrades 1 μg of pUC19 plasmid DNA in 10 minutes at 37°C in standard reaction buffer |
| Sensitivity / LOD | Reduces genomic DNA to below qPCR detection limit from 10 μg of total RNA when used as directed; detects and degrades <1 pg of contaminating plasmid DNA |
| Specificity | Specific for DNA substrates; no detectable RNase activity as measured by incubation with 5 μg MS2 RNA for 4 hours; no detectable protease activity measured with FITC-casein substrate; no exonuclease contamination |
| Reaction Conditions / Protocol | For RNA samples: add 1 U DNase I per 1 μg of RNA in reaction buffer (10 mM Tris-HCl pH 7.5, 2.5 mM MgCl2, 0.5 mM CaCl2), incubate at 37°C for 15–30 minutes, inactivate by adding EDTA to 5 mM final and heating at 65°C for 10 minutes (or 75°C for 5 minutes in presence of EDTA); for DNA footprinting, use optimized buffer conditions as described in protocol |
| Components / Formulation | Lyophilized DNase I (bovine pancreas), 10× reaction buffer (100 mM Tris-HCl pH 7.5, 25 mM MgCl2, 5 mM CaCl2), nuclease-free water for reconstitution; reconstitution to 2 U/μL in 50% glycerol, 20 mM Tris-HCl pH 7.5, 1 mM MgCl2 |
| Storage Conditions | Lyophilized enzyme stored at –20°C (stable for 24 months); reconstituted enzyme stored at –20°C in single-use aliquots (stable for 12 months); avoid repeated freeze-thaw; reaction buffer at –20°C or 2–8°C |
| Shelf Life | 24 months from date of manufacture (lyophilized form) |
| Package Specifications | 10,000 units and 50,000 units; sufficient for processing 10–50 mg of total RNA |
| Product Form | Lyophilized powder with separate reaction buffer |
| Quality Control | Each lot tested: specific activity ≥2,000 Kunitz U/mg; RNase absence verified with radioactive RNA substrate (no degradation in 4 hours); protease absence verified with fluorescent casein; functional DNA removal: 1 μg human genomic DNA degraded below qPCR detection in 15 minutes with 1 U enzyme |
| Key Features | Ultra-pure with no detectable RNase — critical for RNA work; calcium-dependent activity enables controlled reactions with EDTA stop; validated for sensitive qPCR and RNA-seq applications; single-use aliquot format prevents contamination; extensive quality control for absence of contaminating nucleases |
For research use only, not for clinical use.
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