T4 DNA Polymerase,specification,price,image-Bio-Equip in China

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Product Specification

Synonyms	DNA-directed DNA polymerase
Expression System	E.coli
Molecular Weight	105kDa (Reducing)
Purity	＞95% by SDS-PAGE and HPLC
Conjugation	Unconjugated
Tag	His Tag
Physical Appearance	Liquid
Storage Buffer	100 mM K3PO4, 1 mM DTT, 50% Glycerol, pH 6.5 @ 25°C
Stability & Storage	Store at -25 ~ -15℃ for 2 years
Reference	1. Capson TL, Peliska JA, Kaboord BF, Frey MW, Lively C, Dahlberg M, Benkovic SJ. Kinetic characterization of the polymerase and exonuclease activities of the gene 43 protein of bacteriophage T4. Biochemistry. 1992 Nov 17;31(45):10984-94. 2. Tanguy Le Gac N, Delagoutte E, Germain M, Villani G. Inactivation of the 3'-5' exonuclease of the replicative T4 DNA polymerase allows translesion DNA synthesis at an abasic site. J Mol Biol. 2004 Mar 5;336(5):1023-34.

Background

T4 DNA polymerase catalyzes the synthesis of DNA in the 5´→3´ direction and requires the presence of template and primer. This enzyme possesses 3´→5´ exonuclease activity, which is much higher than that found in DNA polymerase I (E. coli). Unlike E. coli DNA polymerase I, T4 DNA polymerase lacks 5’ →3’ exonuclease activity. T4 DNA polymerase can be used to Removal of 3’ overhangs or fill-in of 5’ overhangs to form blunt ends.

Components

3 U/μl T4 DNA Polymerase, 100 mM K3PO4, 1 mM DTT, 50% Glycerol, pH 6.5 @ 25°C
10* Reaction Buffer: 500 mM NaCl, 100 mM Tris-HCl, 100 mM MgCl2, 1000 µg/ml Recombinant Albumin, (pH 7.9 @ 25°C)

Protocol

Incubate the following reaction at 12°C for 15 minutes

Component	Final Concentration
10X Reaction Buffer	1X
dNTP Mix (10 mM)	100 mM each
T4 DNA Polymerase (3,000 U/ml)	1000 U/ml
DNA	10 ug
Nuclease-free Water	to 10 µl
Total Reaction Volume	10 µl

Stop reaction by adding EDTA to a final concentration of 10 mM and heating to 75°C for 20 minutes

Guidelines

Elevated temperatures, excessive amounts of enzyme, failure to supplement with dNTPs or long reaction times will result in recessed ends due to the 3´ → 5´ exonuclease activity of the enzyme.

Unit Definition

One unit is defined as the amount of enzyme that will incorporate 10 nmol of dNTP into acid insoluble material in 30 minutes at 37°C