Nucleic acid amplification is a technique used to produce several copies of a specific segment of RNA/DNA. Amplified RNA and DNA can be used for a variety of applications, such as genotyping, sequencing, and detection of bacteria or viruses. There are two different types of amplification, non-isothermal and isothermal. Non-isothermal amplification produces multiple copies of RNA/DNA through reiterative cycling between different temperatures. Isothermal amplification produces multiple copies of RNA/DNA at a constant reaction temperature. NASBA takes single stranded RNA, anneals primers to it at 65°C, and then amplifies it at 41°C to produce multiple copies of single stranded RNA. In order for successful amplification to occur, an enzyme cocktail containing, Avian Myeloblastosis Reverse Transcriptase (AMV-RT), RNase H, and RNA polymerase is used. AMV-RT synthesizes a complementary DNA strand (cDNA) from the RNA template once the primer is annealed. RNase H then degrades the RNA template and the other primer binds to the cDNA to form double stranded DNA, which RNA polymerase uses to synthesize copies of RNA. One key aspect of NASBA is that the starting material and end product is always single stranded RNA. That being said, it can be used to amplify DNA, but the DNA must be translated into RNA in order for successful amplification to occur.
Loop-mediated isothermal amplification (LAMP) is another isothermal amplification technique.