dsDNase (5000 U)

QIAGEN's dsDNase is a 42.8 kDa recombinant endonuclease derived from marine amphipods and expressed in Pichia pastoris, designed for the rapid and RNA-safe degradation of genomic DNA. The enzyme displays high specific activity towards double-stranded DNA, leaving single-stranded DNA or RNA undamaged under standard conditions, and remains highly active across a broad spectrum of temperatures, buffer conditions, and pH levels. It hydrolyzes phosphodiester linkages to yield oligonucleotides carrying 5′-phosphate and 3′-hydroxyl groups.

The enzyme is supplied in a buffer containing 20 mM Tris-HCl at pH 8.0, 50 mM KCl, 5 mM MgCl2, and 50% (v/v) glycerol. One unit is defined as the amount of enzyme that causes an increase in absorbance at 260 nm of 1.0 in 30 minutes at 37°C and pH 8.0, using herring sperm DNA as substrate. Its specific activity is comparable to bovine DNase I, while dsDNase offers higher stability in demanding reaction and storage conditions such as high-salt and detergent-containing buffers and elevated temperatures. In the presence of Mg2+ or Ca2+ ions, the enzyme shows high endonuclease activity towards dsDNA while leaving ssDNA and RNA intact; Ca2+ ions further increase total activity. Mn2+ ions slightly elevate dsDNase activity but can also act on ssDNA and RNA and should therefore be avoided during RNA or ssDNA purification.

Assay specifications indicate protein purity of greater than 95%, with no RNase activity and no protease activity detected. Quality control procedures include SDS-PAGE with Coomassie Blue detection for protein concentration, gel electrophoresis-based RNase activity assessment using 1 µg of total eukaryotic RNA incubated with 10 U of enzyme in a 20 µL volume for 1 hour at 37°C, and protease activity testing using 1 µg of BSA with 100 U of enzyme for 20 hours at 37°C. For typical digestions, 35 U of dsDNase is recommended for degradation of up to 50 µg of dsDNA at 37°C for 10–15 minutes, though optimal conditions should be determined experimentally. Inactivation is achieved by incubation at 85°C for 15 minutes in the presence of 1–10 mM DTT or TCEP, or alternatively using a spin column or phenol/chloroform extraction.