Meridian is a primary manufacturer of specialized high-quality molecular reagents and offers solutions to a wide range of industries to diagnose and treat diseases, discover new therapeutics or develop tests for environmental, food and cosmetic safety.
Guide to High-Performing Enzymes For PCR, Isothermal amplifications & Next-Generation Sequencing (NGS)
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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
DNA Polymerases
Standard & Hot-Start Taq DNA Polymerase
Taq DNA polymerase is from the thermophilic bacterium Thermus aquaticus and is a heat-stable enzyme commonly used in PCR. Meridian Bioscience’s Taq DNA polymerases are designed for fast and sensitive PCR from complex templates and are available in several hot-start formats including antibody, chemical and aptamer-based technology. The enzymes are supplied with a novel buffer containing dNTPs, MgCl 2 and enhancers that have been optimized for robust amplification and fast cycling conditions, considerably reducing the reaction time without compromising on PCR sensitivity or yield. Hot-start enzymes also ensure that a reaction remains completely inactive during PCR set-up to prevent non-specific amplification.
SPECIFICITY: Highly sensitive and specific amplification using antibody hot-start Taq DNA polymerase to detect down to 1 copy of template
Using antibody hot-start Taq DNA polymerase (MDX008) in Air-Dryable ™ Direct DNA qPCR Blood mix to detect EGFR Exon 19 mutation from 10% human plasma extract. The data demonstrates the specific and sensitive detection of a single copy of EGFR Exon19 deletion amongst 10,000 EGFR Exon19 wildtype copies.
EGFR_EX19 Deletion | EGFR_EX19 Widetype
THERMOSTABILITY: Stable at room temperature for up to one month, streamlining manufacturing processes and enabling convenient shipping and handling
A)
B)
C)
4 ℃ | room temperature
The stability of Taq HS DNA Polymerase (MDX008) is examined in Air-Dryable ™ Direct RNA/DNA qPCR Saliva Mix in a triplex reaction on A) g-actin B) b2mg and C) GAPDH DNA targets at two different concentrations, after being stored at 4°C ( blue ) and room temperature ( red ) for one month. The results illustrate that the stability of the Taq HS DNA polymerase is unaffected, with no loss of functional activity, even at room temperature for one month, facilitating manufacturing process and allowing for easy shipping and handling.
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DNA Polymerases
Glycerol-free Taq DNA Polymerase
Glycerol-free polymerases are ideal for developing ambient-temperature stable assays and offer performance advantages in terms of activity and stability. Meridian’s Glycerol-Free Taq HS polymerase (MDX011) and Glycerol-Free Taq HS High Conc. DNA Polymerase (MDX170) are high-concentration (50 U/ μ L) enzymes optimized to deliver robust amplification without glycerol. The high concentration format of the enzymes is well suited for assay optimization and enzyme titration. In addition, Meridian Glycerol-Free Taq HS DNA Polymerases are compatible with drying technologies such as air-drying or lyophilization. LYO-COMPATIBILTIY: Glycerol-Free Taq Polymerase exhibits the same efficiency and sensitivity after lyophilization
“
Glycerol-Free Hot Start Polymerase is compatible with lyophilization without any loss in sensitivity and speed ”
Wet mix | Lyophilized mix
A 10-fold serial dilution of human DNA (10 3 copies down to 10 copies) using Glycerol-Free Taq HS 50U/ μ L (MDX011) as a wet mix ( black ) or lyophilized mix ( blue ) to illustrate that lyophilization does not have an effect on the quality of the Glycerol-Free Taq HS.
Proof-reading Pfu DNA Polymerase
Pfu is a high-fidelity antibody-mediated hot-start DNA polymerase from the hyperthermophilic archea Pyrococcus furiosus (Pfu). The Pfu polymerase enzyme synthetises DNA in the 5´ → 3´ direction and also possesses a 3´ → 5´ exonuclease (proofreading) activity. Base misincorporations that may occur during polymerization are rapidly excised by this proofreading activity. Meridian’s High- Fidelity Pfu has an error rate of 3.0 x 10 -6 generating blunt-ended amplicons. It can be supplied in a novel buffer system containing dNTPs and enhancers, along with MgCl 2 and is perfect for applications such as target enrichment, NGS library amplification, or cloning. HIGH-FIDELITY: Proof-reading activity of Pfu allows for accurate NGS library amplification.
“
Meridian’s High-Fidelity Pfu exhibits an error rate of 3.0 x 10 -6 ”
Four NGS libraries were created using E. coli DNA. The libraries were amplified using High-Fidelity Pfu (MDX003) prior to being placed on flow cells and sequenced. For each sample, the fraction of uniquely mapped (mapped to only one place on a reference sequence), ambiguously mapped and unmapped reads relative to the total number of reads per sample is shown. The results illustrate that with E. coli DNA, High-Fidelity Pfu allowed for over 90% unique reads that could be aligned to the reference sequence.
DNA Polymerases
Thermostable Tth DNA Polymerase
HIGH-THERMOSTABILITY: Enhances sensitivity for targets with complex secondary structures via high-temp amplification Glycerol-Free HS Tth DNA Polymerase (HC) is a high-concentration thermostable DNA polymerase from Thermus thermophilus with high reverse transcriptase activity in the presence of Mn2+ ions. Its high processivity and reverse transcriptase activity at elevated temperatures allow Glycerol-Free HS Tth DNA Polymerase (HC) to overcome the problems posed by RNA secondary structures. Glycerol-Free HS Tth DNA Polymerase uses antibody-mediated hot start technology that enhances the specificity and sensitivity of PCR. It is supplied with a reaction buffer that contains dNTPs and excipients required for lyophilization. Glycerol-Free HS Tth DNA Polymerase (HC)enables flexible and scalable reaction volumes, and is suitable for applications such as fast RT-qPCR amplification of difficult templates from crude samples.
“
Meridian’s Tth polymerase enables high-efficiency amplification of structurally complex RNA and DNA templates ”
Performance of MDX205 ( red ), MMLV-RT ( blue ) and AMV-RT ( black ) after pre-incubation from 40°C to 95°C for 10 mins, in a multiplex one-step RT-qPCR assay on mouse RNA. ∆ Ct values were calculated against the Ct value produced by the same enzyme stored at -20 °C. The data illustrates that MDX205 maintains its full performance even after 10 min at 95°C, in contrast to MMLV-RT and AMV-RT which are inactivated above 55°C and 60°C, respectively.
Isothermal Enzymes
Glycerol-free Phi29 DNA Polymerase
Glycerol-Free Phi29 DNA Polymerase (HC) is a highly processive DNA-dependent DNA polymerase with strong strand displacement activity, which enables efficient isothermal DNA amplification from low DNA inputs. Glycerol-Free Phi29 DNA Polymerase (HC) high-fidelity amplification, makes it useful for sequencing DNA template preparation, rolling circle amplification (RCA) and multiple displacement amplification (MDA).
AMPLIFICATION POWER: Strong DNA yield even at low template concentrations
“
Glycerol-Free Phi29 is Proven to Deliver Higher Amplification Efficiency ”
DNA polymerase activity of lyophilized Glycerol-Free Phi 29 DNA Polymerase (HC) ( red ) was measured in a rolling circle amplification (RCA) assay, to amplify single stranded circular microphage DNA, and compared against Φ 29 DNA Polymerase (high concentration) from Qiagen ( black ). The results demonstrate that Glycerol-Free Phi 29 DNA Polymerase (HC) has greater DNA polymerization activity than the product from other suppliers.
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Isothermal Enzymes
Bst DNA Polymerase
Bst DNA Polymerase, short for Bacillus stearothermophilus DNA Polymerase, is a type of enzyme commonly used in isothermal DNA amplification methods such as Loop-Mediated Isothermal Amplification (LAMP), Isothermal Helicase-Dependent Amplification (HAD) and Isothermal Multiple Cross Displacement Amplification (MCA). Meridian’s Bst DNA Polymerase exhibits fast 5´- 3´ DNA polymerase activity and strong strand displacement activity to deliver fast amplification speed (time-to-results), yield, salt tolerance, and sensitivity.
SPEED: Bst DNA Polymerase exhibits superior speed and specificity
COMPATIBILTIY: Glycerol-Free Bst Polymerase demonstrates faster time-to-results (TTR) and better reproducibly compared to glycerol containing enzymes
Meridian Bst Polymerase | NEB Bst 2.0 | NEB Bst
Meridian Bst Polymerase | NEB Bst 2.0 | NEB Bst | ArcticZymes
A human pathogenic parasite (Strongyloides stercoralis) was screened using Bst Polymerase ( red ), NEB Bst 2.0 ( purple ) and NEB wild-type Bst Large Fragment ( green ), using the same test conditions across four technical replicates..
CFTR (cystic fibrosis transmembrane receptor) was screened using High Conc. Glycerol-Free Bst ( red ), NEB Bst 2.0 ( purple ), NEB wild-type Bst Large Fragment ( green ) and ArcticZymes IsoPol DNA Polymerase ( blue ), using the same test conditions across four technical replicates..
Nucleic Acid Sequence-Based Amplification (NASBA) is an isothermal amplification technique that uses three enzymes (reverse transcriptase, RNase H, and T7 RNA polymerase) to directly amplify an RNA target sequence at 41°C . Meridian offers a full solution for NASBA enzymes, empowering fast and sensitive isothermal amplification direct from an RNA target. These enzymes are glycerol- free and compatible with lyophilization to develop ambient temperature stable assays, ideal for point-of-care testing. NASBA Enzymes SPEED: High specificity of detection with Meridian’s NASBA enzymes under 1 hour with no detectable NTCs SENSITIVITY: Sensitive detection with Meridian’s NASBA enzymes down to 10 copies of RNA under 1 hour
HIV-1 RNA
0 10 20 30 40
1000 copies 100 copies 1000 copies
100 copies 10 copies
10 copies
HIV-1 RNA | NTC
HIV-1 RNA
Primers and a Molecular Beacon probe were designed for HIV-1 RNA and used in NASBA reactions, with a 1,000 copy, 100 copy and 10 copy serial dilution of the RNA targets.
Primers and a Molecular Beacon probe were designed for HIV-1 RNA and used in a nucleic acid sequence-based amplification (NASBA) reaction using Glycerol-Free T7 RNA Polymerase (HC) (MDX201), Glycerol-Free RNase H (HC) (MDX202) and AMV reverse transcriptase, using 1,000 copies of an HIV-1 RNA target.
Reverse Transcriptases
MMLV-RT
Reverse transcriptases are essential for synthesizing complementary DNA (cDNA) strands from RNA templates. They direct the synthesis of first strand cDNA, which can be used directly as a template for applications such as PCR, qPCR, LAMP, or sequencing workflows. Meridian offers several RTs, each containing unique product features such as high thermostability or lyophilization compatibility for creating room-temperature stable mixes.
One Step RT-qPCR/ RT-LAMP
Two Step RT-qPCR/ RT-LAMP
View our selection chart for details:
Thermostability
Name / Features
Conc.
Cat. No
55C MMLV-RT Compatible with lyophilization or air-drying
MDX117
200 U/ µ L
Up to 60°C
RNase-Tolerant MMLV-RT Contains an RNase inhibitor to improve reaction efficiency
MDX043
Up to 50°C
100x
MMLV-RT Lower RNase H activity
MDX044
100x
Up to 45°C
Lyo-Compatible MMLV-RT Glycerol-free formulation
MDX042
100x
THERMOSTABILITY: Meridian’s 55C MMLV-RT exhibits better thermostablility at temperatures up to 60°C
SENSITIVITY: Meridian’s Lyo-Compatible MMLV-RT allows high sensitivity of low copy number RNA targets
55C MMLV-RT | MMLV-RT | SuperScript III Reverse Transcriptase
Hepatitis virus RNA | NTC
Performance of Meridian’s 55C MMLV-RT ( red , MDX117), standard MMLV-RT ( grey , MDX044) and a SuperScript III Reverse Transcriptase (Thermo, black ) after pre-incubation at 40°C to 70°C for 10 mins, in a multiplex one-step RT-qPCR assay. Δ ct values were calculated against the ct value produced by the same enzyme stored at -20 °C. The data illustrates the increased thermal stability of 55C MMLV-RT when compared to other MMLV-RT enzymes and its ability to efficiently synthesize cDNA at temperatures up to 60°C.
The high efficiency of Lyo-compatible MMLV-RT (MDX042) when used with Lyo-Ready qPCR Mix (MDX021) to detect Hepatitis virus (RNA) allows for high sensitivity detection of low copy number targets (as few as five viral particles).
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Next Generation Sequencing (NGS)
NGS is becoming increasingly accessible, affordable, and user-friendly across research, clinical, and industrial settings. However, most NGS workflows still rely on cold-chain shipping and storage for assay reagents. Meridian’s glycerol-free NGS enzymes support lyophilization, offering a sustainable, ambient-stable alternative. Supplied at high concentration, these enzymes are also ideal for assay miniaturization — a key requirement for point-of-care devices.
Glycerol-Free T4 DNA Polymerase (HC)
A template dependent DNA polymerase that catalyzes the addition of nucleotides to the 3’ end of a DNA strand, extending it in the 5’ to 3’ direction. It also possesses 3’ to 5’ exonuclease activity but lacks a 5´to 3´ exonuclease activity. In NGS library preparation it can be used separately or as part of an End-Repair Mix, where it can blunt DNA ends of double-stranded DNA fragments for subsequent ligation of adaptors.
No loss of activity after lyophilization
Lyophilized Glycerol-Free T4 DNA Polymerase (HC) ( red ) was incubated at 37°C for 1 month and its DNA polymerization activity tested against a non-lyophilized aliquot stored at -20°C ( blue ). The results show that the lyophilized Glycerol-Free T4 DNA Polymerase (HC) is stability at ambient temperature for 12 months.
Glycerol-Free DNA Pol I Klenow Fragment (HC)
A large N-terminal truncated protein fragment produced from E. coli DNA polymerase I, which retains polymerase and 3’-5’ exonuclease activity but has lost 5’-3’ exonuclease activity. It can be used to fill in 5´-protruding ends and is ideal for DNA blunting by 3’ overhang removal and fill-in of 5’ overhangs prior to adapter ligation in next-generation sequencing library preparation.
Fast kinetics with high performance
Glycerol-Free DNA Pol I Klenow Fragment (HC) was used in a primer extension assay and compared to other suppliers under the same conditions. The results demonstrate that Glycerol-Free DNA Pol I Klenow Fragment (HC) has a faster reaction rate and higher end fluorescence than most suppliers.
Next Generation Sequencing (NGS)
Glycerol-Free T4 Polynucleotide Kinase (HC)
Used in NGS library preparation, either separately or as part of an End-Repair Mix, where phosphates the 5’-hydroxyl terminus of double stranded DNA for subsequent ligation of adaptors by T4 DNA Ligase.
FAST & EFFICIENT: High-performance enzyme for accelerated workflows
A/ Increase in fluorescence over time and B/ phosphorylation reaction rate of Glycerol-Free T4 Polynucleotide Kinase (HC) ( blue ) and NEB’s T4 Polynucleotide Kinase ( red ), used to demonstrate that the enzymatic activity of Glycerol-Free T4 Polynucleotide Kinase (HC) is comparable to glycerol-containing enzymes from other suppliers.
Glycerol-Free T4 DNA Ligase (HC)
Used to catalyze the formation of a phosphodiester bond between adjacent 5’ phosphate and 3’ hydroxyl groups in duplex DNA, efficiently joining blunt and cohesive ends and repairs single stranded nicks in duplex DNA. Used to join adaptors to DNA, for use in NGS library construction. NUCLEASE-FREE LIGATION: No detectable exo- or endonuclease activity, ensuring optimal ligation efficiency
Glycerol-Free T4 DNA Ligase (HC) was incubated of for 16 hours at 37°C with 1 μ g of HindIII-digested λ DNA (Lanes 2 and 3) and supercoiled pUC19 DNA (Lanes 5 and 6). The results show no detectable exo- and endonuclease activities that may affect the ligation efficiency of the enzyme.
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Custom Enzyme Stabilization Services for Molecular Diagnostics
These services are designed to help you transition from wet chemistry to ambient temperature-stable formats - including lyophilized, air-dried, and liquid-stable assays - without compromising performance. By removing glycerol and applying our formulation expertise, we help preserve enzyme activity while enabling room-temperature storage, simplified logistics, and greater flexibility in assay development and commercialization.
Same enzyme, same protocol – no workflow changes • Improved stability and extended shelf life • Elimination of cold-chain logistics • Simplified manufacturing and faster time to market • Proven performance in infectious disease and oncology applications
Ordering Information
PRODUCT
CAT NO.
Standard and Hot-Start Taq Polymerases Taq DNA Polymerase
MDX001
MDX008
Taq HS DNA Polymerase
MDX015
Aptamer Taq HS (Glycerol-Free)
MDX009
Low DNA Taq HS 5 U/ µ L
Glycerol-free Taq DNA Polymerase Glycerol-Free Taq HS, 50 U/ µ L
MDX011
MDX170
Glycerol-Free Taq HS High Conc. DNA Polymerase
Proof-Reading Pfu DNA Polymerases High-Fidelity Pfu
MDX003
MDX203
Glycerol-Free High-Fidelity Pfu (HC)
MDX205
Glycerol-Free HS Tth DNA Polymerase (HC)
Bst DNA Polymerases Bst DNA Polymerase
MDX012
MDX018
High Conc. Glycerol-Free Bst (100 U/ µ L)
Ordering Information
NASBA Enzymes Glycerol-Free T7 RNA Polymerase (HC, 1,000 U/ µ L)
MDX201
MDX202
Glycerol-Free RNase H (HC, 50 U/ µ L)
MDX209
Glycerol-Free AMV Reverse Transcriptase (HC)
MDX240
Glycerol-Free Phi29 DNA Polymerase (HC)
MMLV-Reverse Transcriptases 55C MMLV-RT
MDX117
MDX042
Lyo-Compatible MMLV-RT
MDX043
RNase Tolerant MMLV-RT
MDX044
MMLV-RT
MDX209
AMV-RT
Next Generation Sequencing (NGS) Glycerol-Free T4 DNA Polymerase (HC)
MDX027
MDX206
Glycerol-Free T4 Polynucleotide Kinase (HC)
MDX208
Glycerol-Free DNA Pol I Klenow Fragment (HC)
MDX200
Glycerol-Free T4 DNA Ligase (HC)
Supporting Products
MDX034 MDX033 MDX075 MDX022 MDX076 MDX007 MDX054 MDX216 MDX056 MDX120 MDX051 MDX067
MDX083 MDX084 MDX063 MDX066 MDX064 MDX065 MDX096 MDX068 MDX069 MDX026 MDX027
1-Step qPCR Buffer, 4x Fast qPCR Buffer, 4x
dNTP Mix, 40mM (Sodium) dNTP Mix, 100mM (Sodium) dATP, 100mM (Sodium) dTTP, 100mM (Sodium) dCTP, 100mM (Sodium) dGTP, 100mM (Sodium) dUTP, 100mM (Sodium)
Inhibitor-Tolerant qPCR Buffer, 5x Lyo-Ready ™ qPCR Buffer, 2.5x
Bst Reaction Buffer, 10x
Taq Dilution Buffer
Uracil DNA Glycosylase* Glycerol-Free UDGase*
VLP-RNA Extraction Control Red VLP-RNA Extraction Control Orange qPCR Extraction Control Red qPCR Extraction Control Orange
RNase Inhibitor
Glycerol-Free RNase Inhibitor dNTP Mix, 100mM (Lithium) dNTP Mix, 10mM (Sodium )
*Not available for sale in the United States
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About Meridian
Meridian is a fully integrated life science company that develops, manufactures, markets, and distributes a broad range of innovative diagnostic products and critical raw materials. We are dedicated to developing and delivering better solutions that give answers with speed, accuracy, and simplicity that redefine the possibilities of life from discovery to diagnosis. As the Life Science division of Meridian, our focus is on supporting immunological and molecular test manufacturers with original raw materials for human, animal, plant, and environmental applications. The large portfolio of antigens, antibodies, blockers, molecular enzymes, nucleotides, and optimized mixes for qPCR and isothermal amplification applications are designed to simplify assay design and enable accurate test results. We strive to provide our customers with solutions they need when they need them – from novel antigens and antibodies to major disease outbreaks such as Zika and SARS-CoV-2 to pioneering the market with our innovative air-dried qPCR/RT-qPCR mixes. We take pride in providing our customers with unparalleled support, customer service, and quality.
Ordering information:
Meridian Life Science, Inc. 5171 Wilfong Road Memphis, TN 38134 Phone: +1 901-382-8716 Fax: +1 901-333-8223 Email: info@meridianlifescience.com Orders: orders@meridianlifescience.com www.meridianbioscience.com/lifescience
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