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                    Products > Routine PCR > Taq DNA Polymerase (P1011/P1012/P1013/P1014/P1015/P1016)

                    Taq DNA Polymerase (P1011/P1012/P1013/P1014/P1015/P1016)


                    Contents (For Research Use Only)

                    Taq DNA PolymeraseCat. NoAmount (U)Note
                    P1011500
                    no dNTPs
                    P1012500contain dNTPs
                    P10131,000no dNTPs
                    P10141,000contain dNTPs
                    P101518,000no dNTPs
                    P101610,000no dNTPs













                    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.

                    Description
                    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.

                    Applications
                    Standard PCR 
                    DNA labeling 
                    DNA sequencing 
                    Numerous applications for which a high-quality, thermostable DNA polymerase is required

                    Unit Definition
                    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.

                    Storage Buffer 


                    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 



                    Note

                    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


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                    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 


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