商品编号


EG-1011



品牌


Excellgen



公司


Excellgen, Inc.



公司分类


DNA Modifying Enzymes



Size

5,000 Units

商品信息



Description


Exonuclease III
(ExoIII) exhibits four catalytic activities. The
3'=>5' exodeoxyribonuclease activity
of ExoIII is specific for double-stranded DNA. ExoIII degrades dsDNA from blunt ends, 5'-overhangs or nicks, releases 5'-mononucleotides from the 3'-ends of DNA strands and produces stretches of single-stranded DNA. A limited number of nucleotides are removed during each binding event, resulting in coordinated progressive deletions within the population of DNA molecules. It is not active on 3'-overhang ends of DNA that are at least four-bases long and do not carry a 3'-terminal C-residue on single-stranded DNA, or on phosphorothioate-linked nucleotides. This property can be exploited to produce unidirectional deletions from a linear molecule with one resistant (3?-overhang) and one suscept
IBL
e (blunt or 5?-overhang) terminus.

ExoIII
3'-phosphatase activity
removes the 3'-terminal phosphate, generating a 3'-OH group. ExoIII
RNase H activity
exonucleolytically degrades the RNA strand in RNA-DNA hybrids. ExoIII
apurinic/apyrimidinic (AP) endonuclease activity
cleaves phosphodiester bonds at apurinic or apyrimidinic sites to produce 5'-termini that are base-free deoxyribose 5'-phosphate residues.




Applications



Creation of unidirectional deletions in DNA fragments in conjunction with S1 Nuclease.

Generation of a single-stranded template for dideoxy-sequencing of DNA.

Site-directed mutagenesis.

Cloning of PCR products.

Preparation of strand-specific probes for dideoxy sequencing





Source

An
E. coli
strain that carries the cloned
E coli

xth
gene



Unit Definition


One unit of the enzyme catalyzes the release of 1 nmol of acid soluble reaction products from
E. coli
[
3
H]-DNA in 30 min at 37°C.




Molecular Weight

31 kDa monomer.



Components



Endonuclease III:

100,000 units/ml in
25 mM Tris-HCl, pH 8.0 @ 25°C, 50 mM KCl, 1.0 mM DTT, 0.1% mM EDTA, 50% Glycerol

10X Reaction Buffer
: 100 mM Tris-HCl, pH 7.0 @ 25°C, 100 mM MgCl
2
, 10 mM DTT.





Quality Control



The absence of endodeoxyribonucleases confirmed by appropriate quality test.

Functionally tested for creation of unidirectional deletions in DNA fragments.





Storage Condition


-20 °C




References



S. G. Rogers, B. Weiss,
Exonuclease III
of Escherichia coli K-12, an AP endonuclease.
Methods Enzymol.

65
, 201-211 (1980).

J. D. Hoheisel, On the Activities of Escherichia coli
Exonuclease III
.
Anal. Biochem.

209
, 238-246 (1993).

S. Henikoff, Unidirectional digestion with
Exonuclease III
creates targeted breakpoints for DNA sequencing.
Gene.

28
, 351-359 (1984).

L-H. Guo, R. Wu, New rapid methods for DNA sequencing based on
Exonuclease III
digestion followed by repair synthesis.
Nucleic Acids Res.

10
, 2065-2084 (1982).

M. A. Vandeyar
et al.
, A simple and rapid method for the selection of oligodeoxynucleotide-directed mutants.
Gene.

65
, 129-133 (1988).

C. Li, R. M. Evans, Ligation independent cloning irrespective of restriction site compatibility.
Nucleic Acids Res.

25
, 4165-4166 (1997).

H. M. Eun,
Enzymology Primer for Recombinant DNA Technology
(Academic Press Inc., San Diego, California, 1996).

J. F. Tomb, G. J. Barcak, Regulating the 3’-5’ activity of
Exonuclease III
by varying the sodium chloride concentration.
BioTechniques.

7
, 932-933 (1989).