How do you use RNase-Free DNase Set?

How do you use RNase-Free DNase Set?

”Prepare DNase I stock solution before using the RNase-Free DNase Set for the first time. Dissolve the lyophilized DNase I (1500 Kunitz units) in 550 μl of the RNasefree water provided. To avoid loss of DNase I, do not open the vial. Inject RNasefree water into the vial using an RNase-free needle and syringe.

What is DNase RNase-free?

DNase I, (RNase-free) is an endonuclease that nonspecifically cleaves DNA to release di-, tri- and oligonucleotide products with 5´-phosphorylated and 3´-hydroxylated ends (1,2). DNase I acts on single- and double-stranded DNA, chromatin and RNA:DNA hybrids.

How do you reconstitute DNase?

Instructions: Add 275 µl DNase/RNase-Free Water to reconstitute the lyophilized DNase I at 1 U/µl. Mix by gentle inversion. Mix by gentle inversion and incubate at room temperature for 15 minutes.

How do you purify RNA after DNase treatment?

After the additional DNase digestion step an additional purification of the RNA from the DNase I enzyme is mandatory. This purification can be done by a cleanup procedure using the High Pure RNA Isolation Kit following the kit protocol (2.2 Isolation of Total RNA from Cultured Cells).

How stable is DNase at room temperature?

It is stable for at least 3 months if stored at room tempera- ture. However, it is recommended to store the DNase I vial at 2 – 8ºC (or below) upon receipt to maintain stability beyond 3 months. Buffer DR and Enzyme Incubation Buffer can be stored at room temperature.

How long can you store DNase?

Store up to 18 months at −15 to −25°C. The solution will not freeze.

What is the difference between DNase and RNAse?

In the laboratory, DNase I is required to remove DNA from samples used in mRNA expression assays, whereas RNase A is used to remove RNA from samples used for DNA analysis. DNase and RNase are important for modifying and metabolizing nucleic acid chains and can be used as disease markers [4–13].

How do you dissolve DNase?

Dissolve 2 mg of crude pancreatic DNase I (Sigma or equivalent) in 1 ml of 50 mM NaCl, Tris-Cl (pH 7.5), 1 mM MgCl2. When the DNase I is dissolved, add 1 ml of glycerol to the solution and mix by gently inverting the closed tube several times. Take care to avoid creating bubbles and foam.

How is DNase contamination removed?

The Standard Technique:

1. Heat at 180°C for at least 8 hours.
2. Rinse in Chloroform.
3. Soak in a 0.1% Aqueous Solution of Diethyl Pyrocarbonate (DEPC) for 2 hours at 37°C.
4. Clean Equipment with a Detergent Solution, rinse thoroughly with Water and Rinse with 95% Ethanol to dry.

Does EDTA inactivate DNase?

EDTA does not inactivate DNase I, it just removes the Mg ion so that the DNase I no longer active due to lack of Mg. You need to add more EDTA than the amount of Mg ions present to inactivate.

What is the QIAGEN RNase-free DNase set?

The QIAGEN RNase-Free DNase Set is guaranteed RNase-free, quality-controlled, and optimized for use with RNeasy procedures and with QIAamp RNA Blood Mini procedures.

Where can I find the DNase-free DNase protocol for RNA digestion?

A protocol for in-solution DNase digestion using the RNase-Free DNase Set can be found in Appendix C of the RNeasy MinElute Cleanup Handbook. For subsequent RNA Cleanup, use either the RNeasy MinElute Cleanup Kit, or follow the instructions for RNA Cleanup in the RNeasy Mini Handbook.

What does RNase-free mean?

The RNase-Free DNase Set provides efficient on-column digestion of DNA during RNA purification using RNeasy Kits or the QIAamp RNA Blood Mini Kit.

How do I get DNase treatment when using the rneasy 96 kit?

For DNase treatment when using the RNeasy 96 Kit, please contact QIAGEN Technical Services or your local distributor for a separate, optimized protocol. The QIAGEN RNase-Free DNase Set is delivered as a stable, lyophilized enzyme. The RNase-Free DNase Set provides 1500 Kunitz units.