Magic in Solution: An Introduction and Brief History of PCR from Carl T. Wittwer and Jared S. Farrar writing in PCR Troubleshooting and Optimization: The Essential Guide : The polymerase chain reaction (PCR) has become a fundamental tool in molecular research and clinical testing. The origins of PCR and its early evolution are described, including adaptation to RNA, thermostable polymerases, automation, improvements in specificity and rapid temperature cycling. Perhaps the most significant advance is real-time PCR, combining both read more ...

High Resolution Melting Analysis from John F. Mackay and Carl T. Wittwer writing in PCR Troubleshooting and Optimization: The Essential Guide : Real-time qPCR using SYBR Green and melting curve analysis to verify specific product amplification has become a standard laboratory technique for rapid, high throughput gene quantification. An extension of this melting curve method - High Resolution melting analysis (HRMA)_ is now doing the same for the analysis of sequence variation, allowing rapid cost-effective discrimination of sequences to SNP read more ...

Difficult Templates and Inhibitors of PCR from Jack M. Gallup writing in PCR Troubleshooting and Optimization: The Essential Guide : One of the least-acknowledged problems with PCR, RT-PCR and qPCR is reaction inhibition. Addressing or eliminating inhibition is central to allowing qPCR to be modeled by the least complex mathematics, and enables more effective troubleshooting of amplifications from difficult templates such as AT- or GC-rich sequences, repetitive sequences, and templates with prohibitive secondary structures. In the absence of read more ...

Cy0 is a new method in Real-time PCR analysis that does not require the assumption of equal efficiency between unknowns and standard curve. The method demonstrates that, in optimal amplification conditions, The Ct, second derivative (Cp), sigmoidal curve fitting method (SCF) and Cy0 methods methods are equally precise and accurate. However, when PCR efficiency was decreased, diluting amplification mix quantity or adding a biological inhibitor such as IgG, the SCF, Ct and Cp methods were markedly impaired while the Cy0 method gave read more ...

I am pleased to provide the following excerpt from a book review of PCR Troubleshooting and Optimization: The Essential Guide : "The book provides a comprehensive selection of the most recently developed applications for PCR use ... This is an essential book for investigators using PCR technology. The PCR-related topics will be of interest to most, if not all, investigators engaged in research that uses this important technique ... a well-balanced book on the many potential uses of this powerful technique. It also provides a number of updated read more ...