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This manual is an indispensable tool for introducing advanced undergraduates and beginning graduate students to the techniques of recombinant DNA technology, or gene cloning and expression. The techniques used in basic research and biotechnology laboratories are covered in detail. Students gain hands-on experience from start to finish in subcloning a gene into an expression vector, through purification of the recombinant protein. The third edition has been completely re-written, with new laboratory exercises and all new illustrations and text, designed for a typical 15-week semester, rather than a 4-week intensive course. The “project approach to experiments was maintained: students still follow a cloning project through to completion, culminating in the purification of recombinant protein. It takes advantage of the enhanced green fluorescent protein - students can actually visualize positive clones following IPTG induction. Cover basic concepts and techniques used in molecular biology research labs Student-tested labs proven successful in a real classroom laboratories Exercises simulate a cloning project that would be performed in a real research lab "Project" approach to experiments gives students an overview of the entire process Prep-list appendix contains necessary recipes and catalog numbers, providing staff with detailed instructions
Advanced Methods in Molecular Biology and Biotechnology: A Practical Lab Manual is a concise reference on common protocols and techniques for advanced molecular biology and biotechnology experimentation. Each chapter focuses on a different method, providing an overview before delving deeper into the procedure in a step-by-step approach. Techniques covered include genomic DNA extraction using cetyl trimethylammonium bromide (CTAB) and chloroform extraction, chromatographic techniques, ELISA, hybridization, gel electrophoresis, dot blot analysis and methods for studying polymerase chain reactions. Laboratory protocols and standard operating procedures for key equipment are also discussed, providing an instructive overview for lab work. This practical guide focuses on the latest advances and innovations in methods for molecular biology and biotechnology investigation, helping researchers and practitioners enhance and advance their own methodologies and take their work to the next level. Explores a wide range of advanced methods that can be applied by researchers in molecular biology and biotechnology Features clear, step-by-step instruction for applying the techniques covered Offers an introduction to laboratory protocols and recommendations for best practice when conducting experimental work, including standard operating procedures for key equipment
The new edition of this popular book emphasizes the decisions that need to be made to select one procedure over another.
This manual is designed as an intensive introduction to the various tools of molecular biology. It introduces all the basic methods of molecular biology including cloning, PCR, Southern (DNA) blotting, Northern (RNA) blotting, Western blotting, DNA sequencing, oligo-directed mutagenesis, and protein expression. Provides well-tested experimental protocols for each technique Lists the reagents and preparation of each experiment separately Contains a complete schedule of experiments and the preparation required Includes study questions at the end of each chapter
The last few years have seen the rapid development of new methodology in the field of molecular biology. New techniques have been regularly introduced and the sensitivity of older techniques greatly improved upon. Developments in the field of genetic engineering in particular have contributed a wide range of new techniques. The purpose of this book therefore is to introduce the reader to a selection of the more advanced analytical and preparative techniques which the editors consider to be frequently used by research workers in the field of molecular biology. In choosing techniques for this book we have obviously had to be selective, and for the sake of brevity a knowledge of certain basic biochemical techniques and terminology has been assumed. However, since many areas of molecular biology are developing at a formidable rate and constantly generating new terminology, a glossary of terms has been included. The techniques chosen for this book are essentially based on those used in a series of workshops on 'techniques in molecular biology' that have been held at The Hatfield Polytechnic in recent years. In choosing these chapters we have taken into account many useful suggestions and observations made by participants at these workshops. Each chapter aims to describe both the theory and relevant practical details for a given technique, and to identify both the potential and limitations of the technique. Each chapter is written by authors who regularly use the technique in their own laboratories.
This volume has evolved from a laboratory methods book that one of us first compiled nearly fifteen years ago. Since that time the book has undergone many minor revisions in order to include new methods and updated versions of older methods. The result has been an increasingly useful and more widely circulated book. However, the recent series of technological explosions generally lumped together under the name of "recombinant DNA technology" has been a turning point in the evolution of this previously underground publication. Minor revisions will no longer do. To keep the book useful we have had to make major revi sions and additions. The result is a dramatically expanded book that should be more useful to more people. The larger size and wider usefulness of the book have made this more formal publication seem a reasonable step to take. One of the reasons that this volume should be useful to many people is that it includes only procedures that have been used repeatedly by us and that have proven highly reliable both to ourselves and to others in our laboratories.
Current techniques for studying biological macromolecules and their interactions are based on the application of physical methods, ranging from classical thermodynamics to more recently developed techniques for the detection and manipulation of single molecules. Reflecting the advances made in biophysics research over the past decade, and now including a new section on medical imaging, this new edition describes the physical methods used in modern biology. All key techniques are covered, including mass spectrometry, hydrodynamics, microscopy and imaging, diffraction and spectroscopy, electron microscopy, molecular dynamics simulations and nuclear magnetic resonance. Each method is explained in detail using examples of real-world applications. Short asides are provided throughout to ensure that explanations are accessible to life scientists, physicists and those with medical backgrounds. The book remains an unparalleled and comprehensive resource for graduate students of biophysics and medical physics in science and medical schools, as well as for research scientists looking for an introduction to techniques from across this interdisciplinary field.
This volume seeks to enable the discovery of tools in chemical biology by providing readers with various techniques ranging from initial chemical genetic screening to target identification. To successfully highlight the essential components of the chemical biology tool discovery process, the book is organizes into four parts that focus on platforms for molecular discovery in in vitro cellular systems, in vivo chemical genetic screening protocols, and methods used to discover functional protein targets. Written in the highly successful Methods of 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. Practical and informative, Chemical Biology: Methods and Protocols seeks to improve the success rate of the chemical biology field through the dissemination of detailed and experiential knowledge.
This laboratory manual gives a thorough introduction to basic techniques. It is the result of practical experience, with each protocol having been used extensively in undergraduate courses or tested in the authors laboratory. In addition to detailed protocols and practical notes, each technique includes an overview of its general importance, the time and expense involved in its application and a description of the theoretical mechanisms of each step. This enables users to design their own modifications or to adapt the method to different systems. Surzycki has been holding undergraduate courses and workshops for many years, during which time he has extensively modified and refined the techniques described here.
The process whereby a single cell, the fertilized egg, develops into an adult has fascinated for centuries. Great progress in understanding that process, h- ever, has been made in the last two decades, when the techniques of molecular biology have become available to developmental biologists. By applying these techniques, the exact nature of many of the interactions responsible for forming the body pattern are now being revealed in detail. Such studies are a large, and it seems ever-expanding, part of most life-science groups. It is at newcomers to this field that this book is primarily aimed. A number of different plants and animals serve as common model org- isms for developmental studies. In Molecular Methods in Developmental Bi- ogy: Xenopus and Zebrafish, a range of the molecular methods applicable to two of these organisms are described, these are the South African clawed frog, Xenopus laevis, and the zebrafish, Brachydanio rerio. The embryos of both of these species develop rapidly and externally, making them particularly suited to investigations of early vertebrate development. However, both Xenopus and zebrafish have their own advantages and disadvantages. Xenopus have large, robust embryos that can be manipulated surgically with ease, but their pseudotetraploidy and long generation time make them unsuitable candidates for genetics. This disadvantage may soon be overcome by using the diploid Xenopus tropicalis, and early experiments are already underway. The transp- ent embryos of zebrafish render them well-suited for in situ hybridization and immunohistochemistry, and good for observing mutations in genetic screens.