Baseline-ZERO™ DNase

Catalogue number

Pack size

DB0711K

1,000MBU

DB0715K

5,000MBU

Baseline-ZERO™ DNase digests double- and single-stranded DNA to mononucleotides more effectively than the commonly used bovine pancreatic DNase I. Even the small DNA oligonucleotides that remain after treatment with bovine pancreatic DNase I are undetectable following treatment with Baseline-ZERO, thus providing a true zero baseline for RNA RT-PCR or microarray gene expression experiments. Complete removal of DNA from RNA preparations is particularly beneficial when RNA in a sample is amplified using a method that involves reverse transcription of RNA using a random primer (e.g., for exon arrays or full coverage expression analysis), since any contaminating DNA would also be a template for random-primed cDNA synthesis.

Baseline-ZERO™ DNase is recommended when you need to be certain that zero DNA remains. Baseline-ZERO™ DNase is available in 1,000 and 5,000 MBU sizes and is suitable for use in each of the following applications.

Last modified: 2008-05-14 16:46:46

Applications

• Removal of genomic DNA from RNA prior to RT-PCR1 (Figure 1) or preparation of target RNA or cDNA for microarray analysis
• Elimination of the DNA template following in vitro RNA synthesis with T7, T3 or SP6 Phage RNA Polymerases
• Removal of ssDNA and dsDNA from viral RNA
• Elimination of genomic DNA from RNA for microinjection and transfection experiments

Baseline-ZERO™ DNase is provided with a 10X reaction buffer and a 10X stop solution. Heat at 65°C for 10 minutes in the stop buffer to inactivate the enzyme activity before reverse transcription.

10X Baseline-ZERO™ DNase Reaction Buffer: 100 mM Tris HCl (pH 7.5), 25 mM MgCl2 and 5 mM CaCl2.

10X Baseline-ZERO™ DNase Stop Solution: 30 mM EDTA.

Unit Definition: One Molecular Biology Unit (MBU) of Baseline-ZERO™ DNase produces an increase in the A260 of a solution of dsDNA, of 0.001 per minute at 25°C. Functionally, 1 MBU completely digests 1µg of pUC19 DNA to mononucleotides in 10 minutes at 37°C.

Storage Buffer: Baseline-ZERO™ DNase is supplied in a 50% glycerol solution containing 50 mM Tris-HCl (pH 7.5), 10 mM CaCl2, 10 mM MgCl2 and 0.1% Triton® X-100.

Quality Control: Baseline-ZERO™ DNase is assayed for its ability to remove intact DNA and oligonucleotides from a preparation of a linear plasmid (Figure 2).

Baseline-ZERO™ DNase is free of detectable RNase activities as assayed by PAGE analysis of 1µg of a synthetic RNA transcript following an overnight incubation with sufficient DNase I to completely digest 1000µg of DNA.

Figure 1
	Figure 1. CT values from real-time PCR of HeLa RNA preparations treated with various DNases. The lower the CT value, the greater the amount of residual DNA not digested by the indicated DNase. Thus, Baseline-ZERO DNase removed all detectable DNA from the RNA sample. The TaqMan probe assay amplified a 268 bp fragment of beta actin. Samples were run in duplicate.
	Figure 2
Figure 2. Demonstration of the use of Baseline-ZERO™ DNase to remove small oligonucleotides during DNase treatment. Lane 1, kilobase ladder; Lanes 2-5, 160 ng of EcoR I-digested plasmid DNA incubated for 15 minutes at 37°C as follows: Lane 2, untreated; Lane 3, incubated with DNase I: Lane 4, with supplier A's hyper-active DNase; Lane 5, with Baseline-ZERO™ DNase. Only Baseline-ZERO™ DNase removes the small residual oligos at the bottom of the gel.

Last modified: 2008-05-14 16:46:46

Protocols for: Baseline-ZERO™ DNase

Baseline-ZERO™ DNase Protocol

(catalogue number DB0711K, DB0715K)

Please note: all protocols off site are the responsibility of the products supplier

Last modified: 2008-05-14 16:46:46

References

1. Kienzle, N. et al., (1996) BioTechniques 20, 612.

Last modified: 2008-05-14 16:46:46