Taq PCR Master Mix Kit
For convenient PCR setup using a premixed solution
For convenient PCR setup using a premixed solution
Cat. No. / ID: 201445
Concentration: 5 units/µl
Recombinant enzyme: Yes
Substrate analogs: dNTP, ddNTP, dUTP, biotin-11-dUTP, DIG-11-dUTP, fluorescent-dNTP/ddNTP
Extension rate: 2–4 kb/min at 72°C
Half-life: 10 min at 97°C; 60 min at 94°C
Amplification efficiency: ≥105 fold
5'–>3' exonuclease activity: Yes
Extra A addition: Yes
3'–>5' exonuclease activity: No
Contaminating nucleases: No
Contaminating RNases: No
Contaminating proteases: No
Self-priming activity: No
The Taq PCR Master Mix Kit includes QIAGEN's Taq DNA Polymerase in a premixed format. This ready-to-use solution also includes the QIAGEN PCR Buffer, MgCl2, and ultrapure dNTPs at optimized concentrations. Only primers and template DNA need to be added to set up PCR. Due to the convenient master mix format, pipetting errors are minimized, ensuring highly reproducible PCR results (see figure " Reproducible PCR"). Taq PCR Master Mix can be stored at 2–8°C for up to 2 months , allowing even faster PCR setup by eliminating thawing time.
TaqDNA Polymerase DNA Polymerase is a high-quality recombinant enzyme that is suitable for general and specialized PCR applications (see figures " Tolerance of different primer Tm values" and " Specific amplification of long PCR products").
The innovative QIAGEN PCR Buffer has been developed to save time and effort by reducing the need for PCR optimization. QIAGEN PCR Buffer contains both KCl and (NH4)2SO4 (see figure " Increased specificity of primer annealing"). This unique buffer facilitates the amplification of specific PCR products. During the annealing step of every PCR cycle, the buffer allows a high ratio of specific-to-nonspecific primer binding. Owing to a uniquely balanced combination of KCl and (NH4)2SO4, the PCR buffer provides stringent primer-annealing conditions over a wider range of annealing temperatures and Mg2+ concentrations than conventional PCR buffers. Optimization of PCR by varying the annealing temperature or the Mg2+ concentration is dramatically reduced and often not required (see figures " Wide annealing temperature window" and " Tolerance to variable magnesium concentration").
The Taq PCR Master Mix Kit is used for standard and specialized applications, including:
|Applications||PCR, RT-PCR, DNA fingerprinting|
|dNTP's included||Yes (in Master Mix)|
|Reaction type||PCR amplification|
|Enzyme activity||5' -> 3' exonuclease activity|
|Real-time or endpoint||Endpoint|
|Sample/target type||Genomic DNA and cDNA|
|Single or multiplex||Single|
|With/without hotstart||Without hotstart|
Both the quality and quantity of nucleic acid starting template affect PCR, in particular the sensitivity and efficiency of amplification. PCR sensitivity and efficiency can be reduced by the presence of impurities in nucleic acid preparations or in biological samples. These PCR inhibitors are completely removed when template is prepared using QIAGEN Kits for nucleic acid purification. Please refer to the Brochure "Maximizing PCR and RT-PCR success" for additional information.
The optimal primer–template ratio has to be determined empirically. If too little template is used, primers may not be able to find their complementary sequences. Too much template may lead to an increase in mispriming events. Generally, no more than 1 ug of template DNA should be used per PCR reaction. As an initial guide, spectrophotometric and molar conversion values for different nucleic acid templates are listed below.
Spectrophotometric conversions for nucleic acid templates
|1 A260 unit*||Concentration (ug/ml)|
*Absorbance at 260 nm = 1
Molar conversions for nucleic acid templates
|1 kb DNA||1000 bp||1.52||9.1 x 1011|
|pUC 19 DNA||2686 bp||0.57||3.4 x 1011|
|pTZ18R DNA||2870 bp||0.54||3.2 x 1011|
|pBluescript II DNA||2961 bp||0.52||3.1 x 1011|
|Lambda DNA||48,502 bp||0.03||1.8 x 1010|
|Average mRNA||1930 nt||1.67||1.0 x 1012|
|Escherichia coli||4.7 x 106*||3.0 x 10-4||1.8 x 108**|
|Drosophila melanogaster||1.4 x 108*||1.1 x 10-5||6.6 x 105**|
|Mus musculus (mouse)||2.7 x 109*||5.7 x 10-7||3.4 x 105**|
|Homo sapiens (human)||3.3 x 109*||4.7 x 10-7||2.8 x 105**|
* Base pairs per haploid genome
** For single-copy genes
PCR products that will be cloned using the QIAGEN PCR Cloning Kit should be generated using a thermostable DNA Polymerase without proofreading activity, such as Taq DNA Polymerase. Such polymerases attach a single A overhang to their reaction products, which can hybridize to the U overhang of the pDrive Cloning Vector. For efficient addition of an A overhang during the PCR procedure, we recommend a final extension step for 10 min at 72°C as described in the standard protocols of the Taq PCR- and HotStarTaq PCR handbook.
Yes. Please see Table 3 in our brochure Maximizing PCR and RT-PCR success. We tested the effects of different inhibitory substances in a number of PCR systems. We also analyzed the effect of including different volumes of reverse transcription (RT) reaction mixtures in PCR. Please see the table below for a list of commonly encountered template impurities and their inhibitory effects on PCR.
Impurities showing inhibitory effects on PCR
|Sodium Acetate||≥5 mM|
|Sodium Chloride||≥25 nM|
|RT reaction mixture||≥15%|
To determine the optimal annealing temperature for a PCR assay, a Temperature Gradient experiment should be performed. To do this, you will set up several PCR reactions in duplicate for the same primer/template combination, using the same PCR chemistry, and subject each of the reactions to a slightly different annealing temperature within a specified range. If a thermal cycler with a temperature gradient function can be used, you can simply program a temperature range for adjacent wells in the cycling block. If no cycler with a gradient function exists in your lab, you will either have to perform duplicate reactions at different temperatures in different machines (if available), or back to back in the same machine.