Thermo Fisher, 70775Y200UN, Sequenase Version 2.0 DNA Polymerase

Thermo Fisher, 70775Y200UN, Sequenase Version 2.
0 DNA Polymerase Description:Sequenase™ Version 2.
0 DNA Polymerase is a genetically engineered form of T7 DNA polymerase. Unlike the wild-type enzyme it has virtually no 3'→5' exonuclease activity. Sequenase Version 2.
0 is highly processive, incorporates nucleotide analogs (dlTP, thio-dNTPs, dideoxy-NTPs, etc.), is not impeded by secondary structures, and can carry out strand displacement synthesis. It is an excellent enzyme for dideoxy-sequencing, and is useful in other applications, especially where the absence of associated exonuclease activity is desirable.
Sequenase Version 2.
0 has two subunits, one is theE. coliprotein thioredoxin (MW 12,000) and the other is a genetically engineered version of bacteriophage T7 gene 5 protein (MW 76,000). The genetic changes in this subunit (a deletion of 28 amino acids accomplished byin vitromutagenesis) eliminate all measurable exonuclease activity without changing the DNA polymerase activity.
Properties:Molecular Weight: Consists of two subunits, modified T7 gene 5 protein (76 kDa) andE. coliThioredoxin (12 kDa)Optimum pH: 7.
5Optimum Temperature: 37°CRequirements for Divalent Cation: Mg2+, Mn2+Purity:Greater than 95% pure as determined by SDS-PAGE.
Tested for contaminating double- and single-stranded endonucleases and exonucleases.
Storage Buffer:20mM potassium phosphate (pH 7.
4), 1mM DTT, 0.
1mM EDTA, 50% glycerol.
Assay Conditions:The reaction mixture (100 μL) contains 40mM Tris-HCl (pH 7.
5), 10mM MgCl2, 5mM DTT, 0.
3mM dNTPs, and 5 μg M13mp18 pre-annealed to 5 pmol M13 universal primer. The enzyme is added to the pre-warmed (37 °C) reaction mixture; incubation is at 37 °C for 1 min.
Unit Definition:One unit of enzyme catalyzes the incorporation of 1 nmol of nucleotide into acid insoluble form in 30 sec at 37 °C.
Concentration:13 units/μLSequenase Dilution Buffer (1 ml included, PN 70765):10mM Tris-HCl (pH 7.
5), 5mM DTT, 0.
1mM EDTA.
References:Tabor, S. and Richardson, C. C. (1989) J. Biol. Chem.
264, 6447-6458.
Wang, D., Coscoy, L., Zylberberg, M., Avila, P. C., Boushey, H. A., Ganem, D. and DeRisi, J. L. (2002)Proc Natl. Acad. Sci. USA,99, 15687-15692.
Paris, M. (1992) Comments 18, (No. 3), United States Biochemical Corporation, Cleveland, Ohio.
Tabor, S. and Richardson, C. C. (1987)Proc.
Natl. Acad Sci. USA84, 4767-4771.
Tabor, S. and Richardson, C. C. (1989)Proc.
Natl. Acad. Sci.
USA86, 4076-4080.
Concentration: 13 units/μL
Description: Sequenase Version 2.
0 DNA Polymerase, 200 Units, 13 units/μL Concentration, Genetically-Engineered form of T7 DNA Polymerase, 7.
5 pH, 37°C Optimum Temperature
For Use With (Application): Sequencing
Polymerase: Genetically-Engineered form of T7 DNA Polymerase
Product Type: Sequenase Version 2.
0 DNA Polymerase
Purity: 0.
95
Quantity: 200 U
Unit Size: Each