In this new edition, the editors have thoroughly updated and dramatically expanded the number of protocols to take advantage of the newest technologies used in all branches of research and clinical medicine today. These proven methods include real time PCR, SNP analysis, nested PCR, direct PCR, and long range PCR. Among the highlights are chapters on genome profiling by SAGE, differential display and chip technologies, the amplification of whole genome DNA by random degenerate oligonucleotide PCR, and the refinement of PCR methods for the analysis of fragmented DNA from fixed tissues. Each fully tested protocol is described in step-by-step detail by an established expert in the field and includes a background introduction outlining the principle behind the technique, equipment and reagent lists, tips on trouble shooting and avoiding known pitfalls, and, where needed, a discussion of the interpretation and use of results.

PCR has been successfully utilized in every facet of basic, cli- cal, and applied studies of the life sciences, and the impact that PCR has had on life science research is already staggering. C- comitant with the essentially universal use of PCR has been the creative and explosive development of a wide range of PCR-based techniques and applications. These increasingly numerous pro- cols have each had the general effect of facilitating and acceler- ing research. Because PCR technology is relatively easy and inexpensive, PCR applications are well within the reach of every research lab. In this sense, PCR has become the "equalizer" between "small" and "big" labs, since its use makes certain projects, especially those related to molecular cloning, now far more feasible for the small lab with a modest budget. This new volume on PCR Protocols does not attempt the impossible task of representing all PCR-based protocols. Rather, it presents a range of protocols, both analytical and preparative, that provide a solid base of knowledge on the use of PCR in many c- mon research problems. The first six chapters provide some basic information on how to get started. Chapters 7-19 represent primarily analytical uses of PCR, both for simple DNA and RNA detection, as well as for more complex analyses of nucleic acid (e. g. , DNA footprin ting, RNA splice site localization). The remaining chapters represent "synthetic," or preparative, uses of PCR.

The correct procedures you need for frustration-free PCR methods and applications are contained in this complete, step-by-step, clearly written, inexpensive manual. - Avoid contamination--with specific instructions on setting up your lab - Avoid cumbersome molecular biological techniques - Discover new applications

Until the mid 1980s, the detection and quantification of a specific mRNA was a difficult task, usually only undertaken by a skilled molecular biologist. With the advent of PCR, it became possible to amplify specific mRNA, after first converting the mRNA to cDNA via reverse transcriptase. The arrival of this technique—termed reverse transcription-PCR (RT-PCR)—meant that mRNA suddenly became amenable to rapid and sensitive analysis, without the need for advanced training in molecular biology. This new accessibility of mRNA, which has been facilitated by the rapid accumulation of sequence data for human mRNAs, means that every biomedical researcher can now include measurement of specific mRNA expression as a routine component of his/her research plans. In view of the ubiquity of the use of standard RT-PCR, the main objective of RT-PCR Protocols is essentially to provide novel, useful applications of RT-PCR. These include some useful adaptations and applications that could be relevant to the wider research community who are already familiar with the basic RT-PCR protocol. For example, a variety of different adaptations are described that have been employed to obtain quantitative data from RT-PCR. Quantitative RT-PCR provides the ability to accurately measure changes/imb- ances in specific mRNA expression between normal and diseased tissues.

PCR Strategies expands and updates the landmark volume PCR Protocols. It is a companion laboratory manual that provides a completely new set of up-to-date strategies and protocols for getting the most from PCR. The editors have organized the book into four sections, focusing on principles, analyses, research applications, and alternative strategies for a wide variety of basic and clinical needs. If you own PCR Protocols, you will want PCR Strategies. If you don't own PCR Protocols, you will want to buy both! - Concepts explained - Methods detailed - Trouble-shooting emphasized - Novel applications highlighted - Key concepts for PCR - Analysis of PCR products - Research applications - Alternative amplification strategies

This second volume focuses on PCR methods and PCR application specificities to the biotechnology and bioengineering field. New and updated chapters detail real-time PCR protocols, synthetic biology applications, pathogen detection, microfluidics, digital, multiplex detection recent advances. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, PCR: Methods and Protocols, Second Edition aims to be a useful and practical guide to new researchers and experts looking to expand their knowledge.

PCR Cloning Protocols, Second Edition, updates and expands Bruce White's best-selling PCR Cloning Protocols (1997) with the newest procedures for DNA cloning and mutagenesis. Here the researcher will find readily reproducible methods for all the major aspects of PCR use, including PCR optimization, computer programs for PCR primer design and analysis, and novel variations for cloning genes of special characteristics or origin, with emphasis on long distance PCR and GC-rich template amplification. Also included are both conventional and novel enzyme-free and restriction site-free procedures to clone PCR products into a range of vectors, as well as state-of-the-art protocols to facilitate DNA mutagenesis and recombination, and to clone the challenging uncharacterized DNA flanking a known DNA fragment.

PCR is the most powerful technique currently used in molecular biology. It enables the scientist to quickly replicate DNA and RNA on the benchtop. From its discovery in the early 80's, PCR has blossomed into a method that enables everything from ready mutation of DNA/RNA to speedy analysis of tens of thousands of nucleotide sequences daily.PCR Applications examines the latest developments in this field. It is the third book in the series, building on the previous publications PCR Protocols and PCR Strategies. The manual discusses techniques that focus on gene discovery, genomics, and DNA array technology, which are contributing factors to the now-occurring bioinformatics boom.Key Features* Focuses on gene discovery, genomics, and DNA array technology* Covers quantitative PCR techniques, including the use of standards and kinetic analysisincludes statistical refinement of primer design parameters* Ilustrates techniques used in microscopic tissue samples, such as single cell PCR, whole cell PCR, laser capture microdissection, and in situ PCREntries provide information on:* Nomenclature* Expression* Sequence analysis* Structure and function* Electrophysiology* Parmacology* Information retrieval

This volume explores and explains how digital PCRs (dPCRs) help in the study of numerous topics, such as infectious diseases, evolution of cancer and treatment responses, somatic mosaicism, genome editing and cell therapy, and food testing for GMOs and pathogens. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and thorough, Digital PCR: Methods and Protocols is a valuable resource for specialists in various fields, including geneticists, neurologists, immunologists, oncologists, and researchers who are interested in environmental sciences.”

Quantitative Real-Time PCR: Methods and Protocols focuses on different applications of qPCR ranging from microbiological detections (both viral and bacterial) to pathological applications. Several chapters deal with quality issues which regard the quality of starting material, the knowledge of the minimal information required to both perform an assay and to set the experimental plan, while the others focus on translational medicine applications that are ordered following an approximate logical order of their medical application. The last part of the book gives you an idea of an emerging digital PCR technique that is a unique qPCR approach for measuring nucleic acid, particularly suited for low level detection and to develop non-invasive diagnosis. Written for the Methods in Molecular Biology series, most chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, laboratory protocols and tips on troubleshooting and avoiding known pitfalls. Practical and authoritative, Quantitative Real-Time PCR: Methods and Protocols aims to aid researchers seeking to devise new qPCR-based approaches related to his or her area of investigation.