How does isothermal amplification work?
Posted May 10, 2024
Isothermal amplification occurs at a constant temperature and involves enzymes to rapidly replicate the target sequence. Isothermal amplification relies on specific DNA polymerases and specialized primer sets to rapidly multiply a target sequence. Various isothermal amplification methods exist, such as loop-mediated isothermal amplification (LAMP), isothermal multiple displacement amplification (IMDA) recombinase polymerase amplification (RPA), and helicase-dependent amplification (HDA). IMDA is a technique that amplifies nucleic acid sequences using a strand-displacing DNA polymerase and multiple primer sets. LAMP involves the use of four to six primers and a strand-displacing DNA polymerase to synthesize DNA rapidly. RPA utilizes a recombinase, primers, a single-stranded DNA binding protein (SSB), and a strand-displacing DNA polymerase. The recombinase (when complexed with the primer) binds with double-stranded DNA at homologous sequences through a strand exchange. Subsequently, a single-stranded binding protein stabilizes the displaced strand and the polymerase extends the primers, generating a new complete copy of the template. HDA utilizes helicase enzymes to unwind the DNA double helix, allowing primer molecules to bind to the exposed single-stranded DNA regions.
Isothermal Amplification Methods for the Detection of Nucleic Acids in Microfluidic Devicesv