Dna Digestion In Crude Rna Extracts And Subsequent Clean Up - Macherey-Nagel NucleoSpin RNA Clean-up XS User Manual

5.2 DNA digestion in crude RNA extracts and subsequent
clean up
Several commonly used RNA purification methods co-purify DNA to a considerable
extent (e.g., phenol based RNA purification). This often requires a subsequent removal of
contaminating DNA and clean up of the RNA from the reaction mixture.
DNA digestion in solution can efficiently destroy contaminating DNA. However, stringent
RNase control and subsequent repurification of the RNA (in order to remove buffer, salts,
DNase, and digested DNA) are usually required.
The MACHEREY-NAGEL rDNase Set (to be ordered separately, see ordering information),
contains high quality, recombinant RNase-free DNase (rDNase) and reaction buffer. It is
optimized for a highly efficient digestion in order to remove even traces of contaminating
DNA.
1 Digest DNA (reaction setup)
Prepare enzyme-buffer premix: Add 1 µL rDNase to 10 µL Reaction Buffer for
rDNase.
Add 1 / 10 volume of enzyme-buffer premix to the crude RNA extract (e.g., to 10 µL
RNA extract add 1 µL of the premix comprising buffer and enzyme).
Gently swirl the tube in order to mix the solutions. Spin down gently (approx. 1 s at
1,000 x g) to collect every droplet of the solution at the bottom of the tube.
Note: Dissolve lyophilized rDNase (rDNase Set, see ordering information) in 540 µL
RNase-free H
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2 Incubate sample
Incubate for 10 min at 37 °C.
3 Repurify RNA
Repurify RNA with the NucleoSpin
NucleoSpin
O as described in the corresponding user manual.
MACHEREY-NAGEL – 03/2021, Rev. 05
®
RNA Clean-up XS
®
RNA Clean up XS kit according to section 5.1.
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