Panning for molecular gold

Birkhauser Boston, 1998. US$125.00 hbk (400 pages)ISBN 0 8176 3945 4In 1848, gold was found on John Sutter’s land and the California Gold Rush started. Rumours of unimaginable riches spread and soon men were in a mad dash to stake claims, hoping that they would be the one to find the ‘golden seam’. Molecular biology spawns similar dreams of finding new genes and answering some of the most important questions in the biomedical sciences.Technological advances over the past 15 years have made molecular biology accessible to biomedical scientists not initially trained in this area. The ongoing sequencing of eukaryotic and prokaryotic genomes, and the power of the polymerase chain reaction (PCR) is attracting researchers into the area of molecular biology with hopes of striking the mother lode. The number of PCR citations escalated from a handful in 1989 to almost 8000 in 1996.The ability to use enzymatic techniques as a molecular microscope into the workings of gene expression is telling us new things about disease, development, and behaviour. However, the task for scientists wishing to harness gene-amplification methods successfully is to understand the powers and the limitations of these approaches. As this field advances rapidly, scientists must be aware of new technological and methodological developments without wasting time on soon-to-be-obsolete technologies.This new book, Gene Quantification, is a welcome addition at this exciting time of intense activity in the field of nucleic-acid quantification. It provides a firm grounding in theoretical aspects of gene quantification, gives concrete examples of current applications and highlights powerful emerging technologies. Nucleic acids exist in vivo in vanishingly small amounts and are therefore inherently difficult to quantify. This compendium centres on technologies that incorporate some form of amplification to enhance signals to readily detectable levels.In his introductory chapter, Ferre makes excellent points about the fundamental characteristics of nucleic-acid quantification. His comments serve as an important reminder of how the traditional experimental concepts of accuracy, precision, and reproducibility must not be forgotten. The very ability to amplify specific genes selectively out of the nucleic ‘soup’ paradoxically makes PCR and other amplification techniques more difficult to use accurately and precisely. Unlike microscopy, where the degree of magnification is known and constant, enzymatic amplification does not have a constant and known degree of ‘molecular magnification’. This uncertainty is compounded by the fact that the amplification, potentially by eight orders of magnitude or more, can amplify errors as well.This book contains insightful chapters on the mathematics of PCR. These chapters give a formulaic basis on which to build quantitative methods, instead of the hand-waving that pervades the field. The mathematical representation of PCR and the application of Poisson statistics to the quantification of RNA and DNA standards provide a firm underpinning, but are unfortunately also challenging reading. Even researchers versed in gene quantification would be well served by revisiting this vital topic.The most exciting chapters deal with new methods and instruments that offer real-time detection of PCR amplification products. Real-time detection assesses product accumulation throughout the progression of the PCR and eliminates traditional post-PCR steps. The simultaneous amplification and detection of nucleic acids reduces the time needed for experiments by as much as 90% while simultaneously increasing accuracy and reproducibility. Several chapters address the two present means of real-time detection, available from Perkin-Elmer/Applied Biosystems and Idaho Technologies/Roche. In their chapter, Wittwer and colleagues present what may well be the future of gene quantification. Utilizing a capillary-based technology, nucleic acids are amplified in 15-sec PCR cycles; products are then discriminated by traditional melting curves (obtained during the denaturation component of the reaction cycle) and the entire experiment takes 30 min. This system offers the potential for easier assessment of multiple genes (multiplex quantification) in one reaction.The chapters on non-PCR-based approaches are an excellent reminder that PCR is not the only gene-quantification method available. These techniques utilize alternative approaches to amplifying nucleic acids or to amplifying the detection signal, and are used primarily in clinical settings. For example, the use of viral load as a tool for monitoring HIV-infection progression (or regression) has been made possible by these amplification methods. Although these systems are primarily used in clinical settings, they have some advantages over conventional PCR and also have great potential for becoming common techniques in all basic scientific research laboratories.This book does have some unrewarding sections where the writing is dry and the content is not significantly different from or better than many current journal articles. These tend to be the examples of specific applications of the quantitative techniques. However, these chapters do not weaken Ferre’s successful illumination of the status of the field and the future potential for gene quantification. Unfortunately, there is not a chapter on ‘gene chips’ or hybrid arrays, an emerging technology of wide interest. This would have been an opportune time to evaluate the strengths and weaknesses of this multiplex approach compared with more traditional amplification technologies. On balance, however, Gene Quantification is a good map through the rapidly changing landscape of nucleic amplification and detection. This book should have a place on many molecular biologists’ shelves and could help an investigator hit a vein of discovery as rich as any found in the California Gold Rush.