PCR Technology: Current Innovations, Third Edition

ISBN-13: 9781138198586 / Angielski / Miękka / 2016 / 457 str.

Tania Nolan; Stephen a. Bustin

Opis

PCR s simplicity as a molecular technique is, in some ways, responsible for the huge amount of innovation that surrounds it, as researchers continually think of new ways to tweak, adapt, and re-formulate concepts and applications. PCR Technology: Current Innovations, Third Edition is a collection of novel methods, insights, and points of view that provides a critical and timely reference point for anyone wishing to use this technology. Topics in this forward-thinking volume include:

• The purification and handling of PCR templates
• The effect of the manufacture and purification of the oligonucleotide on PCR behavior
• Optimum buffer composition
• Probe options
• The design and optimization of qPCR assays
• Issues surrounding the development and refinement of instrumentation
• Effective controls to protect against uncertainties due to reaction variability

Covering all aspects of PCR and real-time PCR, the book contains detailed protocols that make it suitable as both a reference and an instruction manual. Each chapter presents detailed guidelines as well as helpful hints and tips supplied by authors who are recognized experts in their fields. In addition to descriptions of current technology and best practices, the book also provides information about new developments in the PCR arena."

Spis treści

Template Preparation. Pre-PCR Processing Strategies. Use of a Liposomal Internal Control Vehicle for Whole-Process Quality Assurance of Nucleic Acid Amplification. Reaction Components. Oligonucleotide Synthesis and Purification. Augmenting PCR: Effects of Additives in the Polymerase Chain Reaction. Real-Time Fluorescent PCR by Labeled Primer with a Single Fluorescent Molecule. Using Scorpion Primers for Genotyping. LNA: Adding New Functionality to PCR. Modified dNTPs: A Toolbox for Use in PCR. Instruments. Thermocycler Calibration and Analytical Assay Validation. Ultra-High-Speed PCR Instrument Development. Dropletization of Bioreactions: How Single-Cell Quantitative Analyses May Improve Gene Measurement and Cancer Diagnosis. Design, Optimization, QC, and Standardization. Assay Design for Real-Time qPCR. Primer Design for Large-Scale Multiplex PCR and Arrayed Primer Extension. Development and Use of qPCR Assays for Detection and Study of Neglected Tropical and Emerging Infectious Diseases. MIQE: Guidelines for Reliable Design and Transparent Reporting of Real-Time PCR Assays. Data Analysis. Hypothesis-Driven Approaches to Multivariate Analysis of qPCR Data. Experiment Design, Data Management, and Univariate Statistical Analysis of Gene-Expression Data Obtained by Real-Time Quantitative PCR. Biomarker Discovery via RT-qPCR and Bioinformatical Validation. Optimization of Quantitative Real-Time PCR for Studies in Cartilage Mechanobiology. Applications. Enriching DNA Sequences with Nucleotide Variation by Thymidine Glycosylases Combined with Suppression PCR. Chromosome Walking by Inverse PCR. Expression Profiling of MicroRNAs by Quantitative Real-Time PCR: The Good, the Bad, and the Ugly. Dye-Based High-Throughput qPCR in Microfluidic Platform BioMark™. Surface Plasmon Resonance-Based Biosensor Technology for Real-Time Detection of PCR Products. Current Innovations in the Development and Application of the qPCR in a Resource-Limited Setting. A Review of Isothermal Nucleic Acid Amplification Technologies. Proximity Ligation Assay for Protein Quantification. High-Resolution Melt Analysis. Index.