PCR in Studies of AM Fungi: from Primers to Application

Abstract

The polymerase chain reaction (PCR) is an in vitro technique enabling chemical amplification of DNA (Fig. 24.1). With the improvement brought by the use of the heat-stable Taq DNA polymerase of Thermus aquaticus and automation (Saiki et al. 1985; Mullis et al. 1986), it is possible to obtain quick amplification even of single copy genes, starting from minute amounts of material. The impact of this technique in molecular biology is comparable to that which followed the discovery of restriction enzymes. It has been adapted for a wide variety of applications, and in particular PCR has opened the possibility to analyse organisms at the nucleic acid level even when only small amounts of nucleic acid can be obtained, as in the case of arbuscular mycorrhizal (AM) fungi. Furthermore, although the efficiency of PCR amplification is dependent on the purity of the target DNA, Taq DNA polymerase is less sensitive to template purity than other molecular biology techniques, so that partially purified nucleic acid can be used. This feature is a great advantage for plant/soil microbiology research, as investigations can be made directly on partially purified biological material, like fungal spores or infected plant roots.