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Nucleic Acid Sequence Based Amplification (NASBA) is a technique developed in 1991 by J. Compton. It is a sensitive transcription based amplification method. NASBA is especially suited for the amplification of RNA analytes, such as rRNA, mRNA or genomic RNA, as the process entails a reverse transcriptase step. NASBA occurs at low temperatures, prohibiting the amplification of DNA. The process requires the following reagents: reverse transcriptase, RNase H and T7 DNA dependent RNA polymerase, primers targeting the target sequence. NASBA consist of 2 stages: after denaturing the target strand, the first primer, containing a T7 promoter site, binds to the target RNA and is extended by reverse transcriptase into a DNA/RNA hybrid. This RNA/cDNA hybrid is subsequently degraded by RNAse H. In the second step, the second primer binds and forms a double stranded cDNA suitable for amplification. The final result of amplification is many single stranded RNA molecules. Main advantages of NASBA compared to conventional PCR are: Isothermal: NASBA does not require different temperatures Sensitive: NASBA has a similar sensitivity to RT-PCR Product: NASBA’s final product is single stranded RNA, which can easily hybridize to HKLife EFADchips.

NASBA chart



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