Contents (For Research Use Only)
|Taq DNA Polymerase||Cat. No||Amount (U)||Note|
Taq DNA Polymerase has two concentrations with 2.5 U/μl and 5 U/μl, default package is 5 U/μl.
10×PCR Buffer (Mg2+ Plus) can replace with 10×PCR Buffer (Mg2+ free) and 25 mM MgCl2.
Please choose the appropriate package for your experiment.
Taq DNA Polymerase is purified from E. coli. expressing a cloned Thermus aquaticus DNA polymerase gene. This enzyme has a 5' → 3' DNA polymerase and a 5' → 3' exonuclease activity but lacks a 3' → 5' exonuclease activity. The enzyme consists of a single polypeptide with a molecular weight of approximately 94 kDa. Taq DNA polymerase is heat-stable and will synthesize DNA at elevated temperatures from single-stranded templates in the presence of a primer.
Numerous applications for which a high-quality, thermostable DNA polymerase is required
One unit is defined as the amount of the enzyme required to catalyze the incorporation of 10 nM of dNTPs into an acid-insoluble form in 30 minutes at 70°C using hering sperm DNA as substrate.
20mM TrisCl ( pH8.0), 100mM KCl, 3.2mM MgCl2 1mM DTT，0.1% Triton X-100 ,0.1% Tween20, 0.2mg/ml BSA, 50% (v/v) glycerol
10X PCR Buffer with Mg2+
100mM Tris-HCL PH8.8，500mM KCl ，16mM MgCl2 ，1% Triton100
ü Recombinant Taq DNA Polymerase is the enzyme of choice for most PCR applications.
ü The half-life of enzyme is >40 minutes at 95°C.
ü The error rate of Taq DNA Polymerase in PCR is 2.2x10-5 errors per nt per cycle;
the accuracy (an inverse of the error rate) an average number of correct nucleotides incorporated before making an error, is 4.5x104.
ü Taq DNA Polymerase accepts modified nucleotides (e.g. biotin-, digoxigenin-, fluorescent-labeled nucleotides) as substrates for the DNA
synthesis.Compatible with TA cloning – generates PCR products with 3’-dA overhangs.
ü Recommendations with Template DNA in a 50μl reaction volume
6. Incubate for an additional 10 min at 72°C and maintain the reaction at 4°C. The samples can be stored at -20°C until use.
Notes on cycling conditions
ü Initial denaturation can be performed over an interval of 1-5 min at 95℃ depending on the GC content of template.
ü Optimal annealing temperature is 5℃ lower than the melting temperature of primer-temperature DNA duplex. If nonspecific PCR products are obtained optimization of annealing temperature can be performed by increasing temperature stepwise by 1-2℃.
ü The number of PCR cycles depends on the amount of template DNA in the reaction mix and on the expected yield of the PCR product. 25-35 cycles are usually sufficient for the majority PCR reaction. Low amounts of starting template may require 40 cycles.
ü The time of the final extension step can be extended for amplicons that will be cloned into T/A vectors.
7. Analyze the amplification products by agarose gel electrophoresis and visualize by ethidium bromide staining. Use appropriate molecular weight standards.
Store all components at –20°C