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Product Insert
Heat Labile dsDNase, 2 U/µl
LOT: See product label
EXPIRY DATE: See product label
ORDERING INFORMATION
CAT. NO.
SIZE
PACKAGE CONTENT
BR1100801
100 U
(50 rxn)
50 µl Heat Labile dsDNase
BR1100802
500 U
(250 rxn)
5 × 50 µl Heat Labile dsDNase
COMPONENT
COMPOSITION
Heat Labile dsDNase
Heat Labile dsDNase, 2 U/µl, in storage buffer containing 50% (v/v) glycerol.
STORAGE
-20°C (until expiry date – see product label)
FEATURES
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·
·
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Provides endonuclease activity specific for double-stranded DNA
Is easily heat-inactivated at low temperatures
Degrades genomic DNA in RNA preps
Does not affect RNA quality or quantity
APPLICATIONS
·
·
·
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Easy integration into cDNA synthesis protocols
Removal of genomic DNA from RNA preparations
Removal of contaminating DNA from PCR, qPCR and RT-qPCR
Purification of DNA polymerases
PIN-11008-002
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page 1
Heat Labile dsDNase, 2 U/µl
DESCRIPTION
biotechrabbit™ Heat Labile dsDNase is an endonuclease that cleaves phosphodiester bonds in DNA to yield
oligonucleotides with 5'-phosphate and 3'-hydroxyl termini.
The enzyme has a particularly strong preference for double-stranded DNA (dsDNA), thus double-stranded DNA
is degraded, leaving single-stranded DNA and RNA intact. The enzyme is completely and irreversibly inactivated
with 5 min at 55°C in the presence of 1 mM DTT and pH 8.0. Heat Labile dsDNase is highly active at 20–40°C .
PROTOCOL
Prevention of contamination
When assembling the reactions, care should be taken to eliminate the possibility of contamination with undesired
RNA, DNA and nucleases.
·
·
·
·
Use separate clean areas for preparation of samples and reaction mixtures.
Wear fresh gloves. Use sterile tubes and pipette tips with aerosol filters for assay setup.
Use only water and reagents that are free of RNA, DNA and nucleases.
With every assay setup, perform a contamination control reaction that does not include a template.
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Heat Labile dsDNase, 2 U/µl
PROTOCOL FOR REMOVING GENOMIC DNA FROM RNA PREPARATIONS
PROCEDURE
NOTES
· Prepare RNA, eluting with water.
· Avoid elution or dissolution of RNA in TE Buffer,
since EDTA will reduce the activity of the
enzyme.
· Add MgCl2 to a final concentration of 3 mM.
· Add DTT to a final concentration of 1 mM.
· Add your Reaction Buffer of choice depending on the
downstream application. The pH should be kept > 8
and the KCl/NaCl concentration below 50 mM.
· Minimum Mg2+ concentration of 2.5 mM is
necessary for DNase activity.
· Typically, use 2U Heat Labile dsDNase per 50µl
reaction.
· If high pH, salt or other inhibiting factors are present,
amounts of 0.1 U Heat Labile dsDNase for each
microliter of reaction mix should be used.
· Incubate at room temperature for 15 min.
· DTT is necessary for complete inactivation at
55°C.
· DNA is degraded during this step.
· Incubate at 55°C for 5–10 min to inactivate the Heat
Labile dsDNase.
· The DNA-free RNA preparation can be used directly
for reverse transcription in the same tube.
· Store the RNA at 4°C (short-term) or −80°C
(long-term). For long tern storage add 3 mM
EDTA to compensate for the MgCl 2 added
before.
PROTOCOL FOR REMOVING DNA FROM PCR MASTER MIXES
PROCEDURE
NOTES
· Prepare the PCR master mix without template DNA,
including primers, probe (if appropriate) and dNTPs.
· The final reactions must contain no more than
50 mM KCl.
· Heat Labile dsDNase is inhibited by (NH 4)2SO4
and EDTA.
· Add MgCl2 to a final concentration of 3 mM.
· Add DTT to a final concentration of 1 mM.
· Minimum Mg2+ concentration of 2.5 mM is
necessary for DNase activity.
· DTT is necessary for complete inactivation at
55°C.
· Add 1 U Heat Labile dsDNase to each 20–25 µl of final · DNA is degraded during this step.
PCR reaction volume.
· Incubate at 37–40°C for 15 min.
· Incubate at 55°C for 5–10 min to inactivate the Heat
Labile dsDNase.
· Cool down and add the template DNA.
· The decontaminated amplification mix is ready for
PCR.
PIN-11008-002
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page 3
Heat Labile dsDNase, 2 U/µl
CERTIFICATE OF ANALYSIS
Unit Definition
One unit is defined as an increase in absorbance at 260 nm of 0.001 per minute, using 50 mg/ml of high MW DNA
in 50 mM Na-acetate pH 5.0 and 5 mM MgCl 2.
Quality Control
Protein Purity
Purified to apparent homogeneity by SDS-PAGE.
Quality confirmed by: Head of Quality Control
SAFETY INSTRUCTIONS
For safety instructions please see Safety Data Sheets (SDS)/Sicherheitshinweise finden Sie in den SDS unter:
http://www.biotechrabbit.com/support/documentation.html.
USEFUL HINTS
· Visit Applications at www.biotechrabbit.com for more products and product selection guides.
· Most biotechrabbit products are available in custom formulations and bulk amounts.
CONTACT BIOTECHRABBIT
biotechrabbit GmbH
Volmerstr. 9a
12489 Berlin, Germany
[email protected]
[email protected]
www.biotechrabbit.com
Phone: +49 30 555 7821-10
Fax:
+49 30 555 7821-99
Legal Disclaimer and Product Use Limitation
Purchase of product does not include a license to perform any patented applications; therefore it is the sole
responsibility of users to determine whether they may be required to engage a license agreement depending upon
the particular application in which the product is used. This product was developed, manufactured, and sold for in
vitro use only. It is not suitable for administration to humans or animals.
Trademarks: biotechrabbit™ (biotechrabbit GmbH).
valid from 24.08.2016
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PIN-11008-002