| Product |
DeoxyribonucleaseI
(RNase free) - MBI |
| Description |
Deoxyribonuclease I (RNase
free) is a highly purified preparation of bovine pancreatic
deoxyribonuclease I (DNase I). The enzyme is an endonuclease
that digests single- and double-stranded DNA, producing
mono- and oligodeoxyribonucleotides with a 5'-phosphate
and a 3'-hydroxyl groups. DNase I is activated by bivalent
metal ions. In the presence of Mg2+, the enzyme cleaves
each strand of DNA independently, and cleavage sites that
are distributed in a statistically random fashion. In
the presence of Mn2+, DNase I cleaves both strands of
the DNA at approximately the same site yielding fragments
of DNA that are blunt-ended or have protruding termini
of only one or two nucleotides in length. |
| Concentration |
1u/µl |
| Unit
Definition |
One unit of enzyme completely
degrades 1µg of plasmid DNA in 10min at 37°C.
1 unit is equivalent to approximately 1 Kunitz unit |
| Activity
Assay |
40mM Tris-HCl (pH 8.0),
10mM MgSO4, 1mM CaCl2, 1µg of pBR322 DNA |
| 10X
Reaction Buffer |
400mM Tris-HCl (pH 8.0
at 25°C), 100mM MgSO4, 10mM CaCl2 |
| Storage
Buffer |
50mM Tris-COOCH3 (pH 7.5),
10mM CaCl2, 10mM MgCl2 and 50% glycerol |
| Quality
Control |
Tested for the absence
of ribonucleases and for digestion of template DNA following
in vitro synthesis of RNA using T7 phage RNA polymerase |
| Applications |
· Nick-translation
of DNA
· Preparation of DNA-free RNA
·
Elimination of template DNA following in vitro synthesis
of RNA with T7,
T3, SP6 RNA polymerases
· Preparation of DNA-free
RNA prior to RT-PCR
· Studying of DNA-protein
interactions by DNase I footprinting |
| Note |
· Deoxyribonuclease
I can be inactivated by heating at 65°C for 10 minutes
in the presence of 2mM EGTA or EDTA (RNA hydrolyzes during
heating in the absence of chelating agent)
·
The amount of enzyme required to completely digest a given
amount of DNA must be determined empirically |
