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This detailed new edition broadens the scope of the first edition by moving beyond classical display technologies. This book explores methodologies for the generation of natively paired antibody libraries, single cell technologies, alternative scaffolds, and in silico antibody sequence assessments are described. The application of these methods may allow for a generation of improved therapeutics and diagnostic reagents in a shorter time frame. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Genotype Phenotype Coupling: Methods and Protocols, Second Edition serves as an ideal guide for researchers seeking to expand their knowledge of antibody-based therapeutics.
This volume aims at providing state-of-the-art protocols detailing ribosome display, cDNA display, phage display, yeast surface display, and mammalian display. Chapters guide readers through methods and protocols on in vitro methods over prokaryotic display systems, lower eukaryotes, and mammalian cells. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of all necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Genotype Phenotype Coupling aims to provide an overview of current technologies in this exciting and continuously evolving field.
Advances in molecular biological research in the latter half of the twentieth century have made the story of the gene vastly complicated: the more we learn about genes, the less sure we are of what a gene really is. Knowledge about the structure and functioning of genes abounds, but the gene has also become curiously intangible. This collection of essays renews the question: what are genes? Philosophers, historians and working scientists re-evaluate the question in this volume, treating the gene as a focal point of interdisciplinary and international research. It will be of interest to professionals and students in the philosophy and history of science, genetics and molecular biology.
This book represents the results of 45 years of research on a wide range of topics, including atomic physics, single-molecule enzymology, whole-cell metabolism, physiology, pharmacology, linguistics, semiotics, and cosmology. It describes the first comprehensive molecular theory of the genotype-phenotype coupling based on two key theoretical concepts: (i) the conformon, the conformational wave packet in biopolymers carrying both the free energy and genetic information; and (ii) the intracellular dissipative structures, the chemical concentration waves inside the cell that serve as the immediate drivers of all cell functions. Conformons provide the driving forces for all molecular machines in the cell, and intracellular dissipative structures coordinate intra- and intercellular processes such as gene expression and cell-cell communications.One of the predictions made by the cell language theory (CLT) is that there are two forms of genetic information — the Watson-Crick genes transmitting information in time (identified with DNA), and the Prigoginian genes transmitting information in space (identified with RNA expression profiles). The former is analogous to sheet music or written language and the latter is akin to audio music or spoken language, both being coupled by conformons acting as the analog of the pianist. The new theory of DNA structure and function constructed on the basis of CLT can rationally account for most of the puzzling findings recently unearthed by the ENCODE (Encyclopedia of DNA Elements) project.The Cell Language Theory has important applications in biomedical sciences including drug discovery research and personalized medicine on the one hand and in the mind-body research and consciousness studies on the other.
The book presents the first comprehensive molecular theory of the living cell ever published since the cell doctrine was formulated in 1838-1839. It introduces into cell biology over thirty key concepts, principles and laws imported from physics, chemistry, computer science, linguistics, semiotics and philosophy. The author formulates physically, chemically and enzymologically realistic molecular mechanisms to account for basic living processes such as ligand-receptor interactions, enzymic catalysis, force-generating mechanisms in molecular motors, chromatin remodelling, and signal transduction. Possible solutions to basic and practical problems facing contemporary biology and biomedical sciences have been suggested, including pharmacotherapeutics and personalized medicine.
A versatile collection of readily reproducible cell-cell interaction assays for uncovering cellular interactions at the molecular level, both in vitro and in vivo. The protocols cover a diverse set of cell-cell interaction models in both normal and pathological states, are readily adaptable to nearly any cell type and organ system, and include primary data and outcome analysis. In addition, the protocols follow the successful Methods in Molecular BiologyTM series format, each offering step-by-step laboratory instructions, an introduction outlining the principles behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls.
Knowledge of the three-dimensional structure of a protein is absolutely required for the complete understanding of its function. The spatial orientation of amino acids in the active site of an enzyme demonstrates how substrate specificity is defined, and assists the medicinal chemist in the design of s- cific, tight-binding inhibitors. The shape and contour of a protein surface hints at its interaction with other proteins and with its environment. Structural ana- sis of multiprotein complexes helps to define the role and interaction of each individual component, and can predict the consequences of protein mutation or conditions that promote dissociation and rearrangement of the complex. Determining the three-dimensional structure of a protein requires milligram quantities of pure material. Such quantities are required to refine crystallization conditions for X-ray analysis, or to overcome the sensitivity limitations of NMR spectroscopy. Historically, structural determination of proteins was limited to those expressed naturally in large amounts, or derived from a tissue or cell source inexpensive enough to warrant the use of large quantities of cells. H- ever, with the advent of the techniques of modern gene expression, many p- teins that are constitutively expressed in minute amounts can become accessible to large-scale purification and structural analysis.
For over half a century, we have been in the thrall of the double-helicaln structure of DNA, which, in an instant, revealed that information can be transferred between generations by a simple rule, A pairs with T, G pairs with C. In its beautiful simplicity, this structure, along with the table of codons worked out in the following decade, had entranced us into believing that we can fully understand the information content of a DNA sequence, simply by treating it as text that is read in a linear fashion. While we have learned much based on this assumption, there is much we have missed. Far from a passive tape running through a reader, genomes contain information that appears in new forms which create regions with distinct behavior. Some are "gene rich", some mobile, some full of repeats and duplications, some sticking together across long evolutionary distances, some readily breaking apart in tumor cells. Even protein-coding regions can carry additional information, taking advantage of the flexible coding options provided by the degeneracy of the genetic code. The chapters in this volume touch on one or more of three interconnected themes; information can be implied, rather than explicit, in a genome; information can lead to focused and/or regulated changes in nucleotide sequences; information that affects the probability of distinct classes of mutation has implications for evolutionary theory.
With recent studies using genetic, epigenetic, and other molecular and neurochemical approaches, a new era has begun in understanding pathophysiology of suicide. Emerging evidence suggests that neurobiological factors are not only critical in providing potential risk factors but also provide a promising approach to develop more effective treatment and prevention strategies. The Neurobiological Basis of Suicide discusses the most recent findings in suicide neurobiology. Psychological, psychosocial, and cultural factors are important in determining the risk factors for suicide; however, they offer weak prediction and can be of little clinical use. Interestingly, cognitive characteristics are different among depressed suicidal and depressed nonsuicidal subjects, and could be involved in the development of suicidal behavior. The characterization of the neurobiological basis of suicide is in delineating the risk factors associated with suicide. The Neurobiological Basis of Suicide focuses on how and why these neurobiological factors are crucial in the pathogenic mechanisms of suicidal behavior and how these findings can be transformed into potential therapeutic applications.
The first edition of Protein Purification Protocols (1996), edited by Professor Shawn Doonan, rapidly became very successful. Professor Doonan achieved his aims of p- ducing a list of protocols that were invaluable to newcomers in protein purification and of significant benefit to established practitioners. Each chapter was written by an ex- rienced expert in the field. In the intervening time, a number of advances have w- ranted a second edition. However, in attempting to encompass the recent developments in several areas, the intention has been to expand on the original format, retaining the concepts that made the initial edition so successful. This is reflected in the structure of this second edition. I am indebted to Professor Doonan for his involvement in this new edition and the continuity that this brings. Each chapter that appeared in the original volume has been reviewed and updated to reflect advances and bring the topic into the 21st century. In many cases, this reflects new applications or new matrices available from vendors. Many of these have increased the performance and/or scope of the given method. Several new chapters have been introduced, including chapters on all the currently used protein fractionation and ch- matographic techniques. They introduce the theory and background for each method, providing lists of the equipment and reagents required for their successful execution, as well as a detailed description of how each is performed.