Detection of Specific DNA Sequences in Environmental Samples Via Polymerase Chain Reaction

Polymerase Chain Reaction (PCR) amplification can be used to answer many diverse questions relevant to soil microbiology and its application must be guided by the aim of the researcher. PCR allows amplification of specific DNA in vitro. The choice of primer sequences is critical for successful amplification of a specific DNA sequence. A standard amplification protocol initially tests the efficacy of a newly designed and synthesized oligonucleotide primer pair to achieve significant amplification of the desired target sequence. Optimization of PCR amplification of a target sequence using a primer pair is often successfully achieved when purified target sequences are initially employed. There are three recognized methods to identify the PCR reaction products: gel electrophoresis using either an agarose or polyacrylamide gels, detection using specific DNA probes or HPLC separations. Sensitivity of amplification is important when a given DNA sequence is in an environmental sample, but in low concentration or copy number.