Synonyms    Taq Plus DNA Polymerase, Taq Plus DNA, TaqPDNA.

Description    Taq Plus is a mixture of Taq and Pfu. Taq Plus, which is used to improve the reliability and yield of conventional primer extension reaction. Taq Plus has two following advantages over Taq: (1) high fidelity with an error frequency 1.6/10⁶ (or 0.0016/103) during DNA synthesis. (2) Taq Plus increases the efficiency of polymerization reaction, resulting in a great percentage of extenuation reaction completion up to 10 kb to 30 kb. Pfu has a temperature optimum between 72-78℃ and remains > 95% active following 1-hour incubation at 95℃.

Source    Recombinant E.coli contains Thermus aquaticus polymerase gene.

Unit Definition    One unit of the enzyme catalyzes the incorporation of 10nmole of deoxyribonucleotides into a polynucleotide fraction in 30 min at 74℃.

Storage Buffer    20mM Tris-HCl (pH 8.0), 1mM DTT, 0.1mM EDTA, 100mM KCl, Stabilizers, and 50%glycerol.

Concentration    5U/µl.

10X Reaction Buffer    200 mM TrisHCI (pH 8.8)

100 mM KCI

100 mM (NH4)2SO4

20 mM Mg SO4

1% Triton X-100

1 mg/ml bovine serum albumin (BSA).

Note    All reagents, including Taq Plus, should be mixed immediately before use.

Reaction Conditions    DNA synthesis is performed in 100?l of mixture containing 20-200μm dNTPs, 0.3-1μm Promers, 0.1-0.250 mg of template DNA, 10 ?l of 10 x reaction buffer and 2.5-5 units of Taq Plus. Mix the reaction gently, centrifuge briefly and then overlay with light mineral oil. Initially, denature the reaction by incubating at 95? for 5 minutes and then cool to 40-68? for 5 minutes to allow the primers to anneal to the template DNA.

Optimization of DNA Synthesis    It is important to ad the reaction components in the following order1. H₂O.

2. 10 x reaction buffer.

3. dNTPs.

4. DNA template and primers.

5. Taq Plus.

Storage Conditions    Stable for 5 days at 10℃, for longer period of time store at -20℃.