chapter  1
16 Pages

Target Amplification-Based Techniques

WithAntoinette A.T.P. Brink, Peter J.F. Snijders, Chris J.L.M. Meijer

The most commonly used type of target amplification-based techniques are those that rely on the property of deoxyribonucleic acid (DNA) to become single-stranded denatured when heated and double-stranded again re-natured when cooled down. Ligase Chain Reaction uses both a thermo stable DNA polymerase and a DNA ligase. The ligation product is amplified exponentially by cycling in the presence of a second set of adjacent oligonucleotides complementary to the first. Loop-mediated isothermal amplification was developed to overcome the relatively low target specificities of Nucleic Acid Sequence-Based Amplification and strand-displacement amplification inherent to their low incubation temperatures. Direct sequence analysis of amplification products is the confirmation method of choice when the exact sequence of the region flanked by the amplification primers is not known. Direct analysis is a rather insensitive detection method because a relatively large amount of amplification product is necessary to obtain a positive signal.