Selecting the right enzyme is critical to the success of a molecular diagnostic assay.
At Meridian, our team of scientists are dedicated to creating cutting-edge enzymes that maximize efficiency, reliability and accuracy. With a keen focus on precision and performance, we continuously innovate to enable the next generation of molecular assays.
DNA polymerases are used for DNA amplification in PCR (polymerase chain reaction) assays supporting applications such as next-generation sequencing, genotyping, pathogen detection, and quantitation. There are many different types of DNA polymerases, each with different properties and characteristics including thermostability, specificity, processivity, fidelity, proofreading and elongation rate. They are derived from various organisms, such as bacteria, archaea, eukaryotic cells and viruses, evolving over billions of years to perform specialized functions unique to the organism they are derived from. Depending on the type of molecular assay, certain DNA polymerase characteristics are sought after, while others may be less desirable, which is why selecting the right enzyme is essential to developing a successful assay. DNA Polymerases Top 5 critical criteria of DNA polymerases that impact assay performance • SPECIFICITY: Defined as the capacity of a DNA polymerase to accurately and selectively amplify a specific target DNA sequence. Specificity is a critical factor that ensures the amplification process produces the desired DNA product and minimizes the generation of non-specific or off-target products. • PROCESSIVITY: Processivity is defined by the number of nucleotides that can be incorporated during template binding. The level of processivity exhibited by a DNA polymerase is often indicative of its rate of synthesis, speed, and affinity to its substrate molecules. A highly processive DNA polymerase can amplify lengthy DNA templates, sequences characterized by intricate secondary structures, or those with a high GC content. • THERMOSTABILITY: An enzyme’s thermostability refers to its ability to withstand high temperatures and remain functional during the temperature cycling steps of the reaction. If the DNA polymerase denatures or becomes inactive, amplification does not take place. • FIDELITY: Fidelity refers to the ability of a polymerase to insert the correct nucleotide during PCR. High fidelity DNA polymerases possess a proofreading or 3’ → 5’ exonuclease activity that allows them to correct errors made during DNA replication, increasing the accuracy of DNA sequence replication. • COST: Healthcare accessibility relies on affordable and effective diagnostic solutions. Our ability to manufacture DNA polymerases at large scale enables assay developers to meet their cost objectives. As technology and molecular biology techniques advance, new solutions to improve specificity and sensitivity are continually introduced. Hot-start enzymes, which include the use of chemically modified enzymes, antibodies, and aptamers, play a crucial role in minimizing nonspecific amplification and improving PCR specificity sensitivity, by enabling the detection of low-abundance DNA targets. Hot-start enzymes are designed to enhance the specificity and efficiency of DNA amplification and offer greater consistency in PCR reactions, as they ensure that DNA synthesis begins at a specific, controlled temperature. While all hot-start modifications inhibit polymerase activity at room temperature, there are some key differences among them:
Hot-start
Benefits
Limitations
Product MDX009 Low DNA Taq HS
• Renders enzyme inactive at room-temperature • Free of animal-origin components
• Longer activation time at a high temperature for the polymerase to become fully active
Chemical
• Short activation time • Provides a high degree of specificity, and consistent, reproducible results by preventing premature polymerase activity • No activation is required. • Free of animal-origin components • Reversible enzyme activation, allowing the selective release of the enzyme under specific temperature conditions
MDX008 Taq HS DNA Polymerase
• Antibodies may be of animal origin • Higher level of exogenous proteins (i.e., antibodies) present in the reaction • Assembled reactions may not be stable at the benchtop for a long time. • May not work well with primers of low melting temperatures (due to low activation temperature and reversible)
Antibody
MDX015 Aptamer Taq HS (Glycerol- Free)
Aptamer
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