The Methods in Molecular Biology series from Humana Press covers a wide range of topics in molecular biology, focusing on practical methods. RT-PCR Protocols consists of 27 chapters of methods and applications using reverse transcription-PCR (RT-PCR). The contributing authors are mostly from Ireland and Europe, with some contributions from Japan and the US. The editor contributes six chapters. The book is divided into several parts, including “Highly Sensitive Detection and Analysis of mRNA”, “Quantitative RT-PCR”, “In Situ Localization of mRNA Expression”, “RT-PCR in Immunology”, and “RT-PCR in cDNA Cloning”.
The basics of RT-PCR are covered well in several sections, with enough detail to assist the bench scientist. Protocols are presented in detail. The relative merits of priming cDNA synthesis with oligo(dT), random hexamers, and specific primers are provided, although there is less guidance on the use of different reverse transcriptases (MMLV and AMV). The focus is on useful applications of RT-PCR.
Several chapters are particularly interesting. The chapter by Mullan et al. on assessing the genetic heterogeneity of hepatitis C virus provides detailed procedures for amplification and cloning of hepatitis C loci for the study of quasispecies. The chapter by O’Connell on probe generation by RT-PCR is very useful, providing a simple method to generate your own RNA or DNA probes for in situ hybridization and other applications. Mann and Hanski objectively compare RT-PCR with other methods of gene expression analysis (in situ hybridization, Western blot, and immunohistochemistry). The generation of phage display libraries of single-chain antibody sequences is nicely presented by Gruel et al. Finally, the chapter by Healy on cDNA library construction from small quantities of RNA is very useful.
The abbreviation RT-PCR is, in retrospect, unfortunate. These days, RT-PCR can refer to either “reverse transcription” or “real-time” PCR. The focus of this book is on conventional reverse transcription-PCR. Although one chapter is included on real-time quantification of hepatitis C virus using the LightCycler, real-time PCR addicts would be better served by other reference texts or the original scientific literature. In general, RT-PCR succeeds in its goal to provide the reader with useful applications relevant to the “wider research community”.
- © 2003 The American Association for Clinical Chemistry