Endonucleases DNA-specific

HL-dsDNase for removal of gDNA from RNA preparations

Endonucleases DNA-specific

Endonucleases DNA-specific, HL-dsDNase

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HL-dsDNase is especially developed to remove contaminating genomic DNA from RNA preparations. Figure 1 shows that HL-dsDNase can reduce 50 ng of human gDNA to levels non-detectable by qPCR. In figure 2, a human total RNA sample was treated with HL-dsDNase and analysed on the Bio-Rad Experion™ System. The results indicate that HL-dsDNase has minimal impact on RNA quality and quantity.

HL-dsDNase is an engineered version of dsDNase that is rapidly and completely inactivated by incubation for 5 minutes at 58°C with 1 mM DTT at pH 8.0 or above. Chemical inactivation in downstream compatible RT or PCR buffers allows milder heat inactivation and, in some cases, skipping heat inactivation altogether. This makes HL-dsDNase very useful for removal of DNA from RNA preparations since the enzyme may be inactivated using various strategies with reduced risk of auto-degradation of RNA in the presence of magnesium, making HL-dsDNase an ideal enzyme when working with small volumes of RNA.

HL-dsDNase is also an excellent choice for removal of unwanted external DNA from samples prior to analysis of nucleic acids protected by biological membranes, e.g., bacteria, viruses, and sperm. Especially if using downstream analysis methods that might be affected by host cell DNA, such as metagenomic sequencing and STR-profiling. The easy inactivation of HL-dsDNase makes it a fast and efficient alternative to methods like differential extraction, where sample material is often lost.

Recommended applications:

Key advantages with HL-dsDNase:

Double-strand DNA specific endonuclease
Easily heat-inactivated by very moderate heat treatment
High specific activity
Useful for removal of DNA from RNA preps

Figures

Figure 1. HL-dsDNase effectively removes gDNA

HL-dsDNase removes at least 50 ng of gDNA in a 10 µl reaction volume at pH 7.5.

Figure 2. HL-dsDNase treatment has minimal impact on RNA integrity

RNA incubated in buffer (lanes 1-4), water (lanes 5-8) or 0.1 U/µl HL-dsDNase (lane 9-12). Samples were analysed using Bio-Rad Experion™ RNA Eukaryote Total RNA StdSens Assay.

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Figure 3. HL-dsDNase has minimal impact on RNA quantity

HL-dsDNase effectively removes gDNA from RNA without affecting SDHA (mRNA) quantification.

Properties

Unit Definition

One unit is defined as an increase in absorbance at 260 nm of 0.001 per minute, using 50 µg/ml of high MW DNA in 100 mM Na-acetate pH 5.0 and 5 mM MgCl₂.

Recommended Reaction and Conditions

Temp.:20 - 40°C, optimal: 37°C
Salt conc.: (NaCI/KCI): 0 - 50 mM, optimal: 0 mM
Mg²⁺: 4 - 10 mM, minimum: 2.5 mM
pH 7-8.3, optimal: 7.5

Storage Buffer

20 mM Tris-HCl pH 7.5, 2 mM MgCl₂, 10 mM NaCl, 0.01% (v/v) Tween 20^*, 50% (v/v) glycerol^*^*

^*70810 and 70800 contains 0.01% Triton-x-100 instead of Tween20.

^*^*Glycerol-free version does not contain glycerol.

Purity

HL-dsDNase is purified to apparent homogeneity.

RNase Activity

2 U HL-dsDNase was incubated with a 2 kb RNA transcript (1 µg) for 4 hours at 37°C. Agarose gel electrophoresis did not reveal any visible signs of RNA degradation.

Sample Requirement

Purification of nucleic acids prior to HL-dsDNase treatment.

Storage and Stability

HL-dsDNase is stable at -20°C for ≥2.5 years in the supplied storage buffer

Quality Control

Recommended Protocols

Ordering Info

Quantity

Product Name

SKU

Pack Size

Concentration

Price

T7 RNA Polymerase

74100-201

10 000 U

50 U/µl

$ 145.00 USD

Endonucleases DNA-specific