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Isothermal nucleic acid amplification techniques (INAATs) are rapid, cycling-free alternatives to PCR, enabling exponential nucleic acid amplification at constant temperatures. Instead of using high temperatures to separate DNA strands, isothermal amplification relies on DNA polymerases with high strand displacement activity. These methods are widely used in multiple applications, such as rapid diagnoses of plant pathogens and infectious disease agents. INNATs are also particularly valuable in downstream applications such as next-generation sequencing, sanger sequencing, microarray genotyping, SNP genotyping, DNA enrichment, and as templates for cell-free protein expression.
Unlock the power of simplicity with isothermal amplification. Click on below techniques to expand
Loop-mediated isothermal amplification (LAMP)
LAMP is a technique for the amplification of DNA or RNA (when reverse transcriptase is incorporated) based on the formation of stem-loop structures under isothermal conditions. It uses Bst DNA polymerase and a set of four to six specifically designed primers that hybridize to six or eight different parts of the target DNA.
Application notes
- RT-LAMP amplification of clinical SARS-CoV-2 RNA using Invitrogen SuperScript IV RT-LAMP Master Mix
- Real-time LAMP for the detection of human pathogen DNA using SuperScript IV RT-LAMP Master Mix
- Assessing Lyo-ready Bst DNA Polymerase stability after air-drying and lyophilization
- RT-LAMP of potato spindle tuber viroid RNA using SuperScript IV RT-LAMP Master Mix
- Using SuperScript IV RT-LAMP Master Mix to detect DNA from fungal and fungus-like plant pathogens
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Product information materials
More information at thermofisher.com/LAMP
Lyo-ready Bst DNA Polymerase provides exceptionally fast reaction speed with various DNA targets.
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Recombinase polymerase amplification (RPA)
RPA is an isothermal amplification mechanism that operates at low temperature and is based on strand invasion that is accomplished by a cocktail of recombinase enzymes, single-stranded binding proteins, and strand displacing DNA polymerases.
Lyo-ready RPA kit: Detects 1 target among 5 templates
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Rolling circle amplification (RCA)
The process of RCA involves the use of a circular DNA template, usually in the form of a plasmid or a circularized oligonucleotide. As the strand-displacing DNA polymerase moves around the circular template, it continuously synthesizes new copies of DNA, leading to exponential amplification. RCA generates a concatemer containing numerous tandem repeats that are complementary to the circular template.
Application notes
User guide
Lyo-ready Bst DNA Polymerase
RCA reaction yields using Gibson Assembly cloning products as a template. Reaction yields were quantified using the Quant-iT PicoGreen dsDNA Assay Kit. Data represent the mean with standard deviation (n = 3). NTC = nontemplate control
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Multiple displacement amplification (MDA) & Whole genome amplification (WGA)
Multiple displacement amplification (MDA) is the most widely used whole-genome amplification (WGA) technique that utilizes a stranddisplacing DNA polymerase, such as Phi29 polymerase and random hexamers to amplify the genome under isothermal conditions.
EquiPhi29 DNA Polymerase provides superior product yield at higher working temperatures compared to wild-type phi29 DNA polymerase
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Upcoming Webinar. Unlocking the potential of isothermal amplification with RCA and RPA
Learning Objectives
- lUnderstand the potential of isothermal technologies
- lExplore the application of RPA reaction to amplify complex NGS libraries
- lExplore the applications of RCA in enhancing protein profiling and mRNA production, eliminating the necessity for plasmid purification from host organisms.
Webinar. Looking Beyond PCR: Isothermal Amplification
Learn about LAMP and WGA. Limited sample material resulting in insufficient DNA input is a common hurdle for downstream analysis. This problem can be solved using a variety of methods, including techniques that utilize isothermal amplification. In this webinar, we will cover two methods which are commonly used in today’s laboratories: (1) whole genome amplification (WGA) and (2) loop-mediated isothermal amplification (LAMP).
Webinar. Lyophilization: Changing the Game for Molecular Diagnostics
Discover the many apparent (and hidden) benefits of molecular diagnostic assay lyophilization, as well as important considerations in process development. Understand how the challenges of assay lyophilization can be mitigated by incorporating enzymes with low glycerol content that still perform comparably to standard formats
Isothermal amplification selection eBook
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