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Description
MyTaq Blood-PCR Kit offers very fast, highly-specific, direct PCR from a wide range of human and animal whole blood samples, including those preserved with anticoagulants.
Product Highlights
Fast – eliminate complex, slow and costly DNA extraction steps, thereby reducing time to results
Simple – fewer protocol steps greatly reduce the risk of sample loss and contamination and minimizes manual effort
Robust – novel buffer system developed to overcome PCR inhibitors in blood
Sensitive – incorporates MyTaq HS DNA Polymerase that exhibits increased affinity for DNA, thereby improving yield of even the most challenging targets
Flexible – developed for a wide range of blood samples, including EDTA, citrate and heparin preserved samples, thereby avoiding the need for further optimization
Versatile – suitable for a range of PCR applications, including multiplexing, amplification of GC-rich templates and long amplicons
Product Description
MyTaq™ Blood-PCR Kit is recommended for fast, specific and direct PCR from human and animal blood samples. MyTaq Blood-PCR Kit incorporates MyTaq HS DNA Polymerase, the latest generation of very high-performance polymerases unique to Meridian. Furthermore, MyTaq HS has an increased affinity for template DNA, giving a high PCR product yield from the most challenging templates.
The combination of a unique, inhibitor-tolerant buffer system and MyTaq HS DNA Polymerase, ensures the MyTaq Blood-PCR Kit overcomes the PCR inhibitors typically present in blood samples, including anticoagulants (EDTA, citrate and heparin). This leads to significantly increased sensitivity and PCR success rates even with demanding applications such as long amplicons and GC-rich templates In addition to supporting robust PCR amplification, the novel buffer system replaces the need for complicated extraction and purification steps or the use of additives.
The speed and high specificity of MyTaq Blood-PCR Kit makes it highly suited for multiplex PCR and high-throughput genotyping assays. The advanced formulation of MyTaq Blood-PCR Kit allows fast cycling conditions to be used, without compromising PCR specificity and yield.
Applications
SNP genotyping
Genetic testing
Pathogen detection
Blood screening
Paternity testing
MyTaq Blood-PCR Kit and the corresponding kit from Supplier K were used to amplify a 70.9% GC-rich fragment of DNA. Two-fold serial dilutions (Lanes 1-12), prepared from 20% human whole blood in either EDTA (a), lithium heparin (b), sodium heparin (c) or sodium citrate (d) were used as template. The results illustrate that MyTaq Blood-PCR kit gives increased PCR yield at all template concentrations.
MyTaq Blood-PCR Kit was used to amplify a 5 kb fragment of DNA. A two-fold serial dilution, prepared from 20% human whole blood (Lanes 1-12) in lithium heparin, was used as template. HyperLadder 1kb marker (M). The results illustrate that MyTaq Blood-PCR Kit can be used to successfully amplify longer amplicons.
Species-specific primers were used to amplify targets from horse, sheep, cat and dog blood (lanes 1-4). The results illustrate that MyTaq Blood-PCR Kit delivers highly sensitive and specific amplification from blood collected from a broad range of vertebrates. HyperLadder 1kb marker (M).
A 4-plex reaction was run using a two-fold serial prepared from 10% human whole blood in lithium heparin (Lanes 1-4), primers specific for the Phe (237 bp), Myc (450 bp), EGFR (844 bp) and ATP (1.2 kb) gene targets and MyTaq Blood-PCR Kit. HyperLadder 1kb marker (M). The results illustrate that MyTaq Blood-PCR kit has excellent multiplexing capability across a broad range of template concentrations.
I’ve tested a number of products and by far the best one was MyTaq Blood-PCR Kit. The major advantage is that it works on blood samples and crude lysates. This product will also amplify larger amplicons than other products tested, even in a multiplex scenario. Other key advantages include the hot-start and its ability to withstand 30 freeze-thaw cycles.
Markus Zeller, AutoGenomics, California, US
Product Selection
Please refer to the PCR Selection Chart to confirm the recommended product for your PCR application.
MyTaq Blood-PCR Kit can be stored at -20°C. Repeated freeze/thaw cycles should be avoided.
When stored under the recommended conditions and handled correctly, full activity of the kit is retained until the expiry date printed on the outer box label.
All our polymerases are guaranteed for a period of 12 months from the date of purchase. These should be stored at -20°C during this time for optimal retention of activity.
Please Note: We do not recommend the storage of our polymerases at -80 °C as ice crystals could form on the active site, which may affect or destroy the activity of the enzyme.
PCR can be a challenging technique, with various parameters to optimize to achieve the best results. If you are having problems, these could be easily resolved by addressing a few issues. Please see our PCR troubleshooting guide for suggestions and help with your specific problems.
Observation
Recommended Solution(s)
No or low PCR yield
Enzyme concentration too low – increase the amount of enzyme in 0.5 U increments.
Primers degraded – check quality and age of the primers.
Magnesium concentration too low – increase concentration in 0.25 mM increments with a starting concentration of 1.75 mM.
Primer concentration not optimized. Titrate primer concentration (0.3-1 µM); ensuring that both primers have the same concentration.
Template concentration too low – Increase concentration of template.
Perform a positive control to ensure that the enzyme, dNTPs and buffers are not degraded and/or contaminated.
Multiple Bands
Primer annealing temperature too low. Increase annealing temperature. Primer annealing should be at least 5°C below the calculated Tm of primers.
Prepare master mixes on ice or use a heat-activated polymerase.
For problems with low specificity. Try adding 3% DMSO (not supplied) to improve specificity.
Smearing or artifacts
Template concentration too high. Prepare serial dilutions of template.
Too many cycles. Reduce the cycle number by 3-5 to remove non-specific bands.
Enzyme concentration too high - decrease the amount of enzyme in 0.5 U increments.
Extension time too long. Reduce extension time in 0.5-1 minute increments.
During PCR setup at room temperature, standard polymerases will have some activity. When using a hot-start polymerase, this enzyme will have no activity at room temperature, thus reducing the risk of unspecific products in the reaction due to these mis-primed oligonucleotides. This feature makes this type of polymerase ideally suited to high-throughput applications, where the reactions may be left sitting at room temperature for prolonged periods of time.
