RNase-Free DNase Set

用于RNA纯化过程中的DNA酶消化

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RNase-Free DNase Set (50)

Cat. No. / ID: 79254

1500 Kunitz units RNase-free DNase I, RNase-free Buffer RDD, and RNase-free water for 50 RNA minipreps

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Preparations

250

RNase-Free DNase Set 旨在用于分子生物学应用。该产品不能用于疾病诊断、预防和治疗。

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特点

Compatible with RNeasy procedures and the QIAamp RNA Blood Mini Kit
Efficient on-column digestion of DNA during RNA purification

产品详情

QIAGEN RNase-Free DNase Set保证无RNA酶、经严格质控和优化，配合RNeasy操作流程和QIAamp RNA Blood Mini操作流程使用。一般来说，使用RNeasy Kits纯化RNA时，不需要DNA酶消化步骤，因为硅胶膜离心柱技术可有效地去除大部分DNA，而无需DNA酶处理。然而，某些RNA应用对痕量DNA非常敏感，因此需要彻底去除DNA。QIAGEN RNase-Free DNase Set中提供RDD缓冲液，经优化可用于柱上DNA酶消化。该缓冲液也非常适合于溶液中高效DNA酶消化。在使用RNeasy试剂盒和QIAamp RNA Blood Mini Kit从细胞和组织中纯化RNA的过程中，QIAGEN RNase-Free DNase Set提供高效的柱上DNA消化。后续的洗涤步骤可将DNA酶高效去除。当使用RNeasy 96 试剂盒过程中需要DNA酶处理时，请联系QIAGEN Technical Services或当地经销商获取专门优化的实验方案。QIAGEN RNase-Free DNase Set是以稳定的、冻干粉酶形式提供。QIAGEN RNase-Free DNase Set提供1500 Kunitz units。1 Kunitz units是指在25°C、pH 5.0条件下，以高度聚合的DNA为底物，使每毫升底物的A₂₆₀值每分钟升高0.001时所需要的DNase I的量（Kunitz, M. [1950] Crystalline desoxyribonuclease. I. Isolation and general properties, spectrophotometric method for the measurement of desoxyribonuclease activity. II. Digestion of thymus nucleic acid (desoxyribonucleic acid): the kinetics of the reaction. J. Gen. Physiol. 33, 349 and 363）。

绩效

One Kunitz unit is defined as the amount of DNase I that causes an increase in A₂₆₀ of 0.001 per minute per milliliter at 25°C, pH 5.0, with highly polymerized DNA as the substrate (Kunitz, M. [1950] Crystalline desoxyribonuclease. I. Isolation and general properties, spectrophotometric method for the measurement of desoxyribonuclease activity. II. Digestion of thymus nucleic acid (desoxyribonucleic acid): the kinetics of the reaction. J. Gen. Physiol. 33, 349 and 363).

资源

For DNase treatment with QIAGEN or PreAnalytiX RNA purification kits

Download Safety Data Sheets for QIAGEN product components.

Publications

Epithelial organic cation transporters ensure pH-dependent drug absorption in the airway.

Horvath G; Schmid N; Fragoso MA; Schmid A; Conner GE; Salathe M; Wanner A;

Am J Respir Cell Mol Biol; 2006; 36 (1):53-60 2006 Aug 17 PMID:16917073

Chronic ethanol feeding to rats decreases adiponectin secretion by subcutaneous adipocytes.

Chen X; Sebastian BM; Nagy LE;

Am J Physiol Endocrinol Metab; 2006; 292 (2):E621-8 2006 Oct 17 PMID:17047161

Effects of technological processes on the tenacity and inactivation of norovirus genogroup II in experimentally contaminated foods.

Mormann S; Dabisch M; Becker B;

Appl Environ Microbiol; 2009; 76 (2):536-45 2009 Nov 20 PMID:19933338

One-step multiplex real-time PCR assay to analyse the latency patterns of Epstein-Barr virus infection.

Kubota N; Wada K; Ito Y; Shimoyama Y; Nakamura S; Nishiyama Y; Kimura H;

J Virol Methods; 2007; 147 (1):26-36 2007 Sep 17 PMID:17870188

Skp2B stimulates mammary gland development by inhibiting REA, the repressor of the estrogen receptor.

Umanskaya K; Radke S; Chander H; Monardo R; Xu X; Pan ZQ; O'Connell MJ; Germain D;

Mol Cell Biol; 2007; 27 (21):7615-22 2007 Sep 4 PMID:17785450

FAQ

The RNeasy Midi/Maxi Handbook contains a protocol for the use of the RNase-Free DNase Set for on-column DNA digestion on RNeasy midi or maxi spin columns. Incubation times and reagent volumes have been modified from the standard RNeasy Mini procedure. See Appendix E of the RNeasy Midi/Maxi Handbook for the detailed protocol.

FAQ ID -143

Yes, buffer RDD of the RNase-Free DNase Set will still work. Please make sure that the buffer is thawed completely without any precipitates before using it. If precipitates are visible, the buffer should be slightly heated.

-20

Yes. RNeasy MinElute Cleanup Kit can also be used to clean up RNA following DNase digestion, or from crude RNA preparations following organic extraction or alcohol-precipitation.

FAQ ID -429

The exact composition of Buffer RDD is proprietary. Buffer RDD is an important component of the RNase-Free DNase Set, which is used in combination with most RNeasy Kits. The composition and salt concentration of Buffer RDD provides efficient on-column digestion of DNA and also ensures that the RNA remains bound to the column.

FAQ ID -2800

Yes. Even though buffer RDD in the RNase-Free DNase Set is optimized for on-column DNase digestion, the buffer is also well-suited for efficient DNase digestion in solution. Please note that the reaction must be cleaned up after the off-column DNase digest to remove the enzyme and buffer RDD, which will interfere with subsequent RT reactions.

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.

FAQ ID -619

RNeasy 96 Kit procedure requires two RNase-Fee DNase sets (cat. no. 79254) for each 96-well plate.

1. To obtain a concentration of 1.8 Kunitz/µl, dissolve each vial of lyophilized DNase in 833 µl of RNase-free water and combine the reconstituted enzyme solution from both vials into one vial.

2. Take 1 ml of this DNase I solution and mix with 7 ml of Buffer RDD* to prepare a DNase I incubation mix immediately before starting the RNeasy 96 protocol. Vortex briefly and keep on ice until use.

3. Use 80 µl of the DNase I incubation mix (from step 2 above) for each well of the RNeasy 96 plate, to perform on-column DNase digestion, according to instructions in the RNeasy 96 Handbook.

*The RDD Buffer Set for RNeasy 96 is available upon request. Please contact QIAGEN Technical Service for information on ordering. Buffer RDD from QIAGEN is optimized for DNase I digestion on the RNeasy membrane.

FAQ ID -3003

Carry out all procedures in a "DNA-free" workspace (see FAQ 2654). Be sure to include any DNase treatment steps in the recommended RNA isolation procedure or treat RNA separately with RNase-free DNase followed by repurification using a spin-column based method. RNeasy Mini Kit can be easily combined with RNase-free DNase. Alternatively, kits like the RNeasy Plus Universal Tissue already include a DNA removal step. Be sure to double both the units of enzyme and the incubation time recommended by the RNase-free DNase manufacturer. To minimize DNA contamination in your RNA preparations, and avoid the need for supplemental DNase treatments, we recommend using the RT2 First Strand Kit, which includes a highly efficient genomic DNA elimination step before reverse transcription.

NOTE: Our Chemistries are not compatible with AMBION's Turbo DNA-Free Kits.

FAQ ID -2662

To ensure efficient gDNA removal when doing an on-column digest using the RNase-Free DNase Set in combination with RNeasy Mini the following factors are crucial:

prevent overloading by adjusting the amount of starting material to no more than the maximum amounts recommended in the RNeasy Mini Handbook
ensure complete disruption and homogenization of the starting material as instructed in the section 'Disruption and homogenization of starting materials' of the handbook
strictly follow the protocol for on-column DNase Digestion in Appendix D of the RNeasy Mini Handbook (you can let wash buffer RW1 incubate on the column for 3-5 minutes before centrifuging to enhance removal of excess gDNA prior to applying the enzyme)

In the rare case that trace amounts of genomic DNA are still detectable in sensitive downstream applications such as e.g., realtime RT-PCR, an in-solution digest using the RNase-Free DNAase set can be performed. Instructions are presented in Appendix C of the RNeasy MinElute Cleanup Handbook.

Alternatively, a second on-column digest can be carried out in future preparations, immediately following the RW1 wash after the first incubation with DNase.

FAQ ID -1087