
DNA Amplification, PCR & qPCR
As the Polymerase Chain Reaction (PCR) is the most common DNA amplification method in molecular biology, NEB’s product portfolio features a large selection of polymerases geared towards this powerful method. As the first company to sell Taq DNA Polymerase to the research market, the first to discover a PCR-stable, high-fidelity DNA polymerase, and the first to provide reagents for PCR performed in space, NEB has a long history of developing reliable and convenient PCR tools. This commitment has continued with the recent development of OneTaq® DNA Polymerase for robust routine PCR and Q5® High-Fidelity DNA Polymerase for robust, ultra high-fidelity PCR (>280 X Taq fidelity). Both product lines have been developed to tolerate a variety of complex templates without experiencing a loss of performance on high-AT or high-GC targets. A variety of NEB polymerases, including OneTaq, Taq and Q5, also benefit from novel aptamer-based hot start technology that does not require a separate activation step.
For experiments where detection and quantification is required instead of isolation, quantitative PCR (qPCR) uses real-time fluorescence to meaure the amount of a DNA target present at each cycle during a PCR. The most common methods of generating a fluorescent signal are by use of hydrolysis probes (e.g., TaqMan®), or a double-stranded DNA binding dye, (e.g., SYBR® Green). Ideally, qPCR master mixes should be evaluated for high reaction efficiency over a wide linear dynamic range, and low variation between repeated reactions across a broad variety of sample types. NEB’s Luna® product line has been developed to simplify your qPCR reagent selection while accomplishing best-in-class performance.
Despite the ubiquitous nature of PCR and qPCR, it may not be the best option for all amplification needs. For point of care and other diagnostic applications, sequence-specific isothermal amplification methods, that eliminate the need for thermocycling, have been particularly useful. Instead of heat, these methods typically employ a strand-displacing DNA polymerase, like Bst DNA Polymerase, Large Fragment, to separate duplex DNA. To address some of the limitations of current isothermal amplification techniques, NEB has developed the next generation Bst, Bst 2.0 and a WarmStart® version of this enhanced polymerase, which enables room temperature reaction setup, yet is fully active at temperatures greater than 50°C.
RNA molecules can also be detected and manipulated through amplification via the use of reverse transcriptases (RT), which are RNA-dependent DNA Polymerases. RTs polymerize a strand of DNA that is complimentary to the original RNA template and is referred to as cDNA. This cDNA can then be further amplified through PCR, qPCR or isothermal methods as outlined above or detected in a single reaction using one-step RT-qPCR or RT-LAMP.
Nucleic acid amplification is a foundational process in molecular biology and, as a testament to its utility, new protocols and modifications are being developed constantly. At NEB, our goal is to use our understanding of enzymology, and dedication to providing high-quality products, to offer reagents for all of your applications. Please visit the application pages below to learn more.
Despite the ubiquitous nature of PCR and qPCR, it may not be the best option for all amplification needs. For point of care and other diagnostic applications, sequence-specific isothermal amplification methods, that eliminate the need for thermocycling, have been particularly useful. Instead of heat, these methods typically employ a strand-displacing DNA polymerase, like Bst DNA Polymerase, Large Fragment, to separate duplex DNA. To address some of the limitations of current isothermal amplification techniques, NEB has developed the next generation Bst, Bst 2.0 and a WarmStart® version of this enhanced polymerase, which enables room temperature reaction setup, yet is fully active at temperatures greater than 50°C.
RNA molecules can also be detected and manipulated through amplification via the use of reverse transcriptases (RT), which are RNA-dependent DNA Polymerases. RTs polymerize a strand of DNA that is complimentary to the original RNA template and is referred to as cDNA. This cDNA can then be further amplified through PCR, qPCR or isothermal methods as outlined above or detected in a single reaction using one-step RT-qPCR or RT-LAMP.
Nucleic acid amplification is a foundational process in molecular biology and, as a testament to its utility, new protocols and modifications are being developed constantly. At NEB, our goal is to use our understanding of enzymology, and dedication to providing high-quality products, to offer reagents for all of your applications. Please visit the application pages below to learn more.
TaqMan® is a registered trademark of Roche Molecular Systems, Inc.
SYBR® is a registered trademark of Molecular Probes, Inc.
Choose Type:
DNA Amplification, PCR & qPCR includes these areas of focus:
- Isothermal Amplification
- PCR
- Routine PCR
- High-Fidelity PCR
- PCR & Reaction Cleanup
- Polymerases for DNA Manipulation
- qPCR & RT-qPCR
- Dye-based qPCR & RT-qPCR
- Probe-based qPCR & RT-qPCR
- RT-PCR & cDNA Synthesis
- cDNA Synthesis
- RT-PCR
- Site Directed Mutagenesis
- Specialty PCR
- Extraction-Free PCR
- Hot Start PCR
- Long Range PCR
- Fast PCR
- Multiplex PCR
- Bisulfite Sequencing
- Polymerases for NGS Library Preparation
- Whole Genome Amplification
FAQs for DNA Amplification, PCR & qPCR
Protocols for DNA Amplification, PCR & qPCR
- PCR Using NEBNext® High-Fidelity 2X PCR Master Mix (M0541)
- PCR Using Q5® Hot Start High-Fidelity DNA Polymerase (M0493)
- Protocol for Q5® Hot Start High-Fidelity 2X Master Mix
- Protocol for Q5® High-Fidelity 2X Master Mix
- PCR Optimization (E0555)
- Protocol for a Routine PCR (E0555)
- PCR Using Q5® High-Fidelity DNA Polymerase (M0491)
- Protocol for a PCR reaction using NEBNext® Q5® Hot Start HiFi PCR Master Mix (M0543)
- Luna® Universal qPCR Master Mix Protocol (#M3003)
- Luna® Universal One-Step RT-qPCR Kit Protocol (E3005)
- Luna® Universal Probe One-Step RT-qPCR Kit Protocol (E3006)
- Luna® Universal Probe qPCR Master Mix Protocol (M3004)
- PCR Brochure
- Polbase
- DNA Polymerase Selection Chart
- Luna® One-Step RT-qPCR Troubleshooting Guide
- Luna® qPCR Troubleshooting Guide
- PCR Troubleshooting Guide
- Choosing RNA Input Amounts for NEB T2010
- General Guidelines for Successful RNA Purification Using the Monarch Total RNA Miniprep Kit
- Guidelines for PCR Optimization with OneTaq® and OneTaq® Hot Start DNA Polymerases
- Guidelines for PCR Optimization with Taq DNA Polymerase
- Guidelines for PCR Optimization with Thermophilic DNA Polymerases
- Optimization Tips for Luna® One-Step RT-qPCR
- Optimization Tips for Luna® qPCR
- A Single-tube, Low Input Protocol for Long Read Sequencing (2019)
- Amplification Reagents for Molecular Diagnostics Applications (2017)
- Colorimetric LAMP: Visual Detection for Simple Diagnostics (2017)
- Molecular Diagnostics for Gastrointestinal Parasites and Impact on Intestinal Microbiota in Rural Agentinian Children (2015)
Tools & Resources
Brochures
Web Tools
Selection Tools
Troubleshooting Guides
Usage Guidelines
Posters
- Gardner, A.F., Jackson, K.M., Boyle, M.M., Buss, J.A., Potapov, V., Gehring, A.M., Zatopek, K.M., Correa, I.R., Jr., Ong, J.L., Jack. W.E. 2019. Therminator DNA polymerase: Modified nucleotides and unnatural substrates Front Mol Biosci. 6, PubMedID: 31069234, DOI: 10.3389/fmolb.2019.00028
Publications related to DNA Amplification, PCR & qPCR
Legal Information
This product is covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB).
While NEB develops and validates its products for various applications, the use of this product may require the buyer to obtain additional third party intellectual property rights for certain applications.
For more information about commercial rights, please contact NEB's Global Business Development team at [email protected].
This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
While NEB develops and validates its products for various applications, the use of this product may require the buyer to obtain additional third party intellectual property rights for certain applications.
For more information about commercial rights, please contact NEB's Global Business Development team at [email protected].
This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.