ABSTRACT
The development of molecular methods to detect and analyze microorganisms from the natural environment without prior culture has provided fascinating insights into microbial diversity and function. Much of this work has been facilitated by the use of the polymerase chain reaction (PCR) to amplify sequences from DNA and RNA extracted directly from environmental samples (see Chapters 1, 2 and 5) (1, 2). However, a major drawback of microbial ecological studies based on classical end-point PCR is that the process inherently introduces biases as the mixed environmental target template is amplified. However, the resulting PCR amplicons, although providing a rich source of information relating to previously uncharacterized genes, cannot be used accurately to quantify numbers or proportions of specific genes or phylotypes within natural environments.
