Historically, the major emphasis on the study of purinergic systems has been predominantly in the areas of physiology and gross pharmacology. The last decade has seen an exponential in- crease in the number of publications related to the role of both adenosine and A TP in mammalian tissue function, a level of interest that has evolved from a more molecular focus on the identity of adenosine and A TP receptor subtypes and the search for selective ligands and development of radioligand binding assays by Fred Bruns and colleagues (especially that for A receptors) that played z a highly significant role in advancing research in the area. In the 60 years since adenosine was first shown to be a potent hypotensive agent, a considerable investment has been made by several pharmaceutical companies-including Abbott, Byk Gulden, Takeda, Warner-LambertlParke Davis, Boehringer Mann- heim, Boehringer Ingelheim, Nelson/Whitby Research and CffiA- Geigy-as well as John Daly's laboratory at the National Institutes of Health, to design new adenosine receptor ligands, and both agon- ists and antagonists with the aim of developing new therapeutic entiities. Numerous research tools have derived from these efforts including: 2-chloroadenosine, R-PIA (~-phenylisopropyladeno- sine; NECA (5' N-ethylcarboxamidoadenosine); CV1808; CI936; PD 125,944; ~-benzyladenosine; PACPX; CPX; CPT; XAC; CGS 15943 and CGS 21680. Yet in the realm of therapeutics it was only in 1989 that adenosine itself was approved for human use in the treatment of supraventricular arrythmias.

This second edition provides updated and expanded chapters covering a broad sampling of useful and current methods in the rapidly developing and expanding field of bioinformatics. Bioinformatics, Volume I: Data, Sequence Analysis, and Evolution, Second Edition is comprised of three sections: Data and Databases, Sequence Analysis, and Phylogenetics and Evolution. The first section details bioinformatics methodologies in the generation of sequence and structural data and its organization into conceptual categories, and databases to facilitate further analyses. The Sequence Analysis section describes the fundamental methodologies for processing the sequences of biological molecules: techniques that are used in almost every pipeline of bioinformatics analysis, particularly in the preliminary stages of such pipelines. Last but not least, the phylogenetics and evolution section deals with methodologies that compare biological sequences for the purpose of understanding how they evolved. As a volume in the highly successful Methods in Molecular Biology series, chapters feature the kind of detail and expert implementation advice to ensure positive results. Comprehensive and practical, Bioinformatics, Volume I: Data, Sequence Analysis, and Evolution, Second Edition is an essential resource for graduate students, early career researchers, and others who are in the process of integrating new bioinformatics methods into their research.

Understanding seed-related processes is of major social, environmental, and economic concern. The viability and vigor of seeds are the very basis for sustainable agriculture and forestry, and comprehending the molecular and cellular events underlying these processes will become increasingly important to many economical sectors and for species that provide the world's food supply. Seed Dormancy: Methods and Protocols covers analytical methods and approaches which have already lead to significant advances in the understanding of seed dormancy and germination. Chapters cover explanations of processes leading to the induction, maintenance, and termination of seed dormancy, the classification of different dormancy types, as well as an overview of protocols used for dormancy-termination of seeds of conifer species. This volume emphasizes methods essential for abscisic acid (ABA) analyses, including methods that have been important for receptor identification, analyses of ABA-catabolizing enzymes (the 8'-hydroxylases), and identification of novel signal transduction components, interacting partners, and/or response factors. The volume closes by addressing the development of new technologies, including spectroscopic methods (some of which allow for non-destructive sampling) as well as highly effective tissue-printing methods for seed dormancy research. Written in the successful Methods in Molecular Biology (TM) series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Seed Dormancy: Methods and Protocols features detailed methods that will prove invaluable for both applied and fundamental seed research.

A comprehensive, state-of-the-art review of the biochemistry, molecular and cellular biology, pharmacology, physiology, and pathology of natriuretic peptides. Written by the foremost researchers in the field, the reviews range from describing the actions of these peptides on the kidney, adrenal gland, vasculature, and brain, to their effects on immune regulation, reproduction, and cell growth. Special attention is given to the potential involvement of these hormones in clinical pathology and to their possible therapeutic use. Especially enlightening is the first-time review of molecular understanding of key cellular signaling pathways and of the importance of the vascular smooth muscle-endothelial interface.

Mass spectrometry underwent dramatic changes during the decade of the 1980s. Fast atom bombardment (F AB) ionization, developed by Barber and coworkers, made it possible for all mass spectrometry laboratories to analyze polar, highly functionalized organic molecules, and in some cases ionic, inorganic, and organometallic compounds. The emphasis of much of this work was on molecular weight determination. Parallel with the development of ionization methods (molecular weight mass spectrometry) for polar biological molecules, the increased mass range of sector and quadrupole mass spectrometers and the development of new instruments for tandem mass spectrometry fostered a new era in structural mass spectrometry. It was during this same period that new instrument technologies, such as Fourier transform ion cyclotron resonance, radio frequency quadrupole ion trap, and new types of time-of-flight mass spectrometers, began to emerge as useful analytical instruments. In addi tion, laser methods useful for both sample ionization and activation became commonplace in almost every analytical mass spectrometry laboratory. In the last 5 years, there has been explosive growth in the area of biological mass spectrometry. Such ionization methods as electrospray and matrix-assisted laser desorption ionization (MALDI) have opened new frontiers for both molecular weight and structural mass spectrometry, with mass spectrometry being used for analysis at the picomole and even femto mole levels. In ideal cases, subfemtomole sample levels can be successfully analyzed. Sample-handling methods are now the limiting factor in analyz ing trace amounts of biological samples."

This volume is a collection of the contributions presented at the 42nd Erice Crystallographic Course whose main objective was to train the younger generation on advanced methods and techniques for examining structural and dynamic aspects of biological macromolecules. The papers review the techniques used to study protein assemblies and their dynamics, including X-ray diffraction and scattering, electron cryo-electron microscopy, electro nanospray mass spectrometry, NMR, protein docking and molecular dynamics. A key theme throughout the book is the dependence of modern structural science on multiple experimental and computational techniques, and it is the development of these techniques and their integration that will take us forward in the future.

The health of the proteome depends upon protein quality control to regulate the proper synthesis, folding, translocation, and clearance of proteins. The cell is challenged constantly by environmental and physiological stress, aging, and the chronic expressions of disease associated misfolded proteins. Substantial evidence supports the hypothesis that the expression of damaged proteins initiates a cascade of molecular events that leads to Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, Huntington's disease, and other diseases of protein conformation.

Animal cell technology is a growing discipline of cell biology which aims to understand the structure, function and behaviour of differentiated animal cells, and especially the development of such abilities as are useful for industrial purposes. These developments range from clonal expansion of differentiated cells with useful abilities, to optimization of cell culture on industrial scale and modulation of the cells' abilities to produce drugs and monoclonal antibodies. The sixth volume in this series gives a complete review of today's state of the art in Japan, a country where this field is especially well advanced. It will be of interest to cell biologists, biochemists, molecular biologists, immunologists and other disciplines related to animal cell culture, working in the academic environment as well as in (biotechnology or pharmaceutical) industry.

At some point in their careers, virtually every scientist and technician, as well as many medical professionals, regardless of their area of specialization have a need to utilize cell culture systems. Updating and significantly expanding upon the previous editions, Basic Cell Culture Protocols, Fourth Edition provides the novice cell culturist with sufficient information to perform the basic techniques, to ensure the health and identity of their cell lines, and to be able to isolate and culture specialized primary cell types. The intent of this extensive volume is to generate a valuable resource containing clear methodologies pertinent to current areas of investigation, rather than attempting to educate cell culturists on specific cell types or organ systems. It is written in the highly successful Methods in Molecular Biology trademark], 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. Comprehensive and up-to-date, Basic Cell Culture Protocols, Fourth Edition compiles the essential techniques needed to approach this vital laboratory activity with full success.

As one of the fastest growing fields of research in the 21st century, nanotechnology is sure to have an enormous impact on many aspects of our lives. Nanostructure Design: Methods and Protocols serves as a major reference for theoretical and experimental considerations in the design of biological and bio-inspired building blocks, the physical characterization of the formed structures, and the development of their technical applications. The chapters contributed by leading experts are divided into two sections, the first of which covers experimental aspects of nanostructure design and the second delves into computational methods. As a volume of the highly successful Methods in Molecular Biology (TM) series, this collection pulls together cutting-edge protocols, written in a step-by-step, readily reproducible format certain to guide researchers to the desired results. Comprehensive and essential, Nanostructure Design: Methods and Protocols uses biological principles and vehicles on design to aid scientists in the great challenges still ahead.

This important volume is mainly concerned with the development of methods for "sequencing" - that is, determination of the order of the amino acids in proteins and of nucleotides in RNA and DNA. In 1943 the position of only one amino acid in a protein (insulin) was known, and Sanger's first paper resulted in finding a second amino acid. In his final paper in 1982 he describes the determination of a DNA sequence of 48,502 nucleotides. The papers describe the steady improvements in techniques, and exciting biological results revealed by the sequences.

Molecular networks provide descriptions of the organization of various biological processes, including cellular signaling, metabolism, and genetic regulation. Knowledge on molecular networks is commonly used for systems level analysis of biological function; research and method development in this area has grown tremendously in the past few years. This book will provide a detailed review of existing knowledge on the functional characterization of biological networks. In 15 chapters authored by an international group of prolific systems biology and bioinformatics researchers, it will organize, conceptualize, and summarize the existing core of research results and computational methods on understanding biological function from a network perspective.

Plant Cell and Tissue Culture continues the high standards of Humana's Methods in Molecular Biology series. Its step-by-step approach (a hallmark of the series) is applied to a wide range of basic laboratory techniques and culture conditions appropriate to plant cells. Because of the diversity of cell types, species, and culture methods, much of this volume is devoted to the culture of particular cell types and to the regeneration of these cells into whole plants. Special attention is also given to the genetic modification of plants, as well as to the economic significance of plant products. Chapters cover a wide range of topics and techniques, including:* tissue culture media and selection * cryopreservation * callus culture techniques * organ culture * embryogenesis * batch culture * large-scale culture * hormonal control * fertilization techniques * gene transfer * cell immobilization * production systems * cell product purification * DNA expression * electrofusion of plant cells * mutant selection * mutagenesis techniques * automation * transfer of nuclei * protoplast culture * media analysis * micropropagation. A detailed appendix lists the formulas for the most commonly employed plant cell media. Comprehensive, easy to follow, and a pleasure to use, Pollard and Walker's Plant Cell and Tissue Culture is an essential tool for everyone--at all levels of proficiency and experience--involved in plant culture.

Examining the chemical modification of biological polymers and the emerging applications of this technology, Chemical Modification of Biological Polymers reflects the change in emphasis in this subsection of biotechnology from the study of protein structure and function toward applications in therapeutics and diagnostics.

Highlights

• The basic organic chemistry of the modification proteins, nucleic acids, oligosaccharides, polysaccharides, and their applications
• New analytical technologies used to characterize the chemical modification of biological polymers
• Identification of in vivo, non-enzymatic chemical modification of biological polymers
• Specific chemical modifications to generate biopharmaceutical products

This book covers the basics on the organic chemistry underlying the chemical modification of biopolymers, including updates on the use of various chemical reagents. It describes the current status of chemical modification of biological polymers and emerging applications of this technology in biotechnology. These technologies are important for the manufacture of conjugate proteins used in drug delivery, for the preparation of nucleic acid microarrays, and for the preparation of hydrogels and other materials used in tissue engineering.

The aim of this volume is to provide a comprehensive overview of optical tweezers setups, both in practical and theoretical terms, to help biophysicists, biochemists, and cell biologists to build and calibrate their own instruments and to perform force measurements on mechanoenzymes both in isolation in vitro and in living cells. Chapters have been divided in three parts focusing on theory and practical design of optical tweezers, detailed protocols for performing force measurements on single DNA- and microtubule/actin-associated mechanoenzymes in isolation, and describing recent advances that have opened up quantitative force measurements in living cells. 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 cutting-edge, Optical Tweezers: Methods and Protocols aims help to further expand the accessibility and use of optical traps by scientists of diverse disciplines.

The filamentous fungi are perhaps unique in the diversity of their metabolic activities. This includes biosynthetic as well as degradative activities, many is to provide an of which are of industrial interest. The objective of this text up-to-date and broad review which emphasizes the genetic and molecular biological contribution in the field of fungal biotechnology. This text begins with an overview of the tools and methodologies involved which, to a large extent, have been developed in the model filamentous fungus Aspergillus nidulans and subsequently have been extended to commercially important fungi. This is followed by a chapter which provides a compilation of genes isolated from commercial fungi and their present status with respect to structure, function and regulation. Chapters 3 and 4 highlight the degradative powers of filamentous fungi. First, a discussion of what is known regarding the molecular genetics of fungi and the genes and enzymes involved in the beverage and food industries. This has an oriental flavour, reflecting the tremendous importance of fungi in traditional Chinese and Japanese food production. An account of lignocellulose degradation by filamentous fungi follows, illustrating the potential of fungi to utilize this substance as a renewable energy source. The ability of fungi to produce high-value foreign proteins is reviewed in chapters 5 and 6. Chymosin production, in particular, represents a good example of high-level yields being obtained, such as to warrant commercial production.

This volume introduces the reader to the latest experimental and bioinformatics methods for DNA sequencing, RNA sequencing, cell-free tumour DNA sequencing, single cell sequencing, single-cell proteomics and metabolomics. Chapters detail advanced analysis methods, such as Genome-Wide Association Studies (GWAS), machine learning, reconstruction and analysis of gene regulatory networks and differential coexpression network analysis, and gave a practical guide for how to choose and use the right algorithm or software to handle specific high throughput data or multi-omics data. 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 cutting-edge, Computational Systems Biology: Methods and Protocols aims to ensure successful results in the further study of this vital field.

As imaging technologies and approaches have evolved, the scope of certain imaging techniques has moved far beyond the production of purely illustrative images or appealing time-lapse movies to providing the scientist with a rich range of ways to measure and quantify the biological process and outcome of gene expression. In Imaging Gene Expression: Methods and Protocols, expert authors offer up-to-date approaches and protocols that scientists in the field have developed, which would benefit the broader scientific community. Divided in three convenient parts, this detailed book covers the output of a gene, namely the RNA molecules that are transcribed from the gene and the way by which these molecules can be tracked or quantified in fixed or living cells, protocols that focus on the gene, DNA, or chromatin, as well as a variety of ways by which nuclear processes intertwined with gene expression can be followed and quantified in living cells as well as approaches for studying several sub-nuclear structures found in eukaryotic cells. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective subjects, lists of materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and up-to-date, Imaging Gene Expression: Methods and Protocols will serve researchers working toward imaging in the context of complete organisms.

Aimed at research scientists and biotechnologists, this book is an essential reading for those working with extremophiles and their potential biotechnological application. Here, we provide a comprehensive and reliable source of information on the recent advances and challenges in different aspects of the theme. Written in an accessible language, the book is also a recommended as reference text for anyone interested in this thriving field of research. Over the last decades, the study of extremophiles has provided ground breaking discoveries that challenge our understanding of biochemistry and molecular biology. In the applied side, extremophiles and their enzymes have spawned a multibillion dollar biotechnology industry, with applications spanning biomedical, pharmaceutical, industrial, environmental, and agricultural sectors. Taq DNA polymerase (which was isolated from Thermus aquaticus from a geothermal spring in Yellowstone National Park) is the most well-known example of the potential biotechnological application of extremophiles and their biomolecules. Indeed, the application of extremophiles and their biologically active compounds has opened a new era in biotechnology. However, despite the latest advances, we are just in the beginning of exploring the biotechnological potentials of extremophiles.

Since the publication of the first edition, lentivirus vector-based technologies, through in vitro and in vivo gene transfer in eukaryotic animal cells, continue to offer the most promising opportunities for curing genetic disorders, as well as cancer and infectious diseases. Lentivirus Gene Engineering Protocols, Second Edition reflects the spectacular progress made in the field with a set of cutting-edge methods contributed by highly respected scientists. Beginning with a thorough overview of the most recent lentivirus developments, the book continues with detailed protocols including sections on the advances in lentiviral vector technology, new lentiviral vector applications, involving transgenic human embryonic stem cells and fetal gene therapy among other topics, as well as the invaluable breakthroughs in LV-mediated expression of microRNAs. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective subjects, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and notes sections, highlighting tips on troubleshooting and avoiding known pitfalls.

Authoritative and timely, Lentivirus Gene Engineering Protocols, Second Edition covers the most relevant issues and techniques of LV-based gene engineering, thus representing a complete theoretical and practical guide for scientists still unfamiliar with LV technologies and those who simply wish to know more about this vital area of study.

This first volume of the comprehensive, two-volume work on oxidative stress in lung disease introduces the molecular mechanisms, and the role of oxidants in the progression of different lung diseases. The lungs of humans and animals are under constant threat from oxidants from either endogenous (e.g. in situ metabolic reactions) or exogenous sources (e.g. air pollutants). Further, oxidative stress causes the oxidation of proteins, DNA and lipids, which in turn generates secondary metabolic products. The book consists of sections, each focusing on different aspects of oxidant-mediated lung diseases. As such it is a unique reference resource for postgraduate students, biomedical researchers and also for the clinicians who are interested in studying and understanding oxidant-mediated lung diseases. The second volume will incorporate other aspects of oxidant-mediated lung diseases, including prevention and therapeutics.

Since the first edition of this book dedicated to differential display (DD) technology was published in 1997, we have witnessed an explosive interest in studying differential gene expression. The gene-hunting euphoria was initially powered by the invention of DD, which was gradually overtaken by DNA microarray technology in recent years. Then why is there still the need for second edition of this DD book? First of all, DD still enjoys a substantial lead over DNA microarrays in the ISI citation data (see Table 1), despite the h- dreds of millions of dollars spent each year on arrays. This may come as a surprise to many, but to us it implies that many of the DNA microarray studies went unpublished owing to their unfulfilled promises (1). Second, unlike DNA microarrays, DD is an "open"-ended gene discovery method that does not depend on prior genome sequence information of the organism being studied. As such, DD is applicable to the study of all living organisms-from bacteria, fungi, insects, fish, plants, to mammals-even when their genomes are not sequenced. Second, DD is more accessible technically and financially to most cost-conscious "cottage-industry" academic laboratories. So clearly DD still has its unique place in the modern molecular biological toolbox for gene expression analysis.

Busy clinicians and health practitioners recognize the importance of speedy detection of pathogens to impede the further spread of infection, and to ensure their patients' rapid and complete recovery. This reader-friendly reference is a unique collection of the newest and most effective diagnostic techniques currently in use in clinical and research laboratories. Instructive commentary regarding the application of these often complex methods is provided. This essential text aids readers in selecting the most efficient method, finding the necessary resources, and avoiding the most common pitfalls in implementation.

The study of animal viruses contributes to the general understanding of the molecular basis of viral infection. The emergence of the SARS (Severe Acute Respiratory Syndrome) virus in the human population, coming from an animal source, highlights the importance of animals in harboring infectious agents. In addition, it has been recognized recently that influenza viruses, which persist in their natural avian host, can directly infect humans. In this book, an international panel of leading virologists provide a state-of-the-art overview of the field, comprehensively detailing the current understanding of viruses, their replication, evolution, and interaction with the host. The authors emphasize strategic and methodological aspects of current research, and provide key related references. Topics include foot-and-mouth disease virus, Pestivirus, Arteriviridae, Coronaviruses (including SARS), Herpesviridae, Paramyxoviridae, influenza viruses, Reoviridae, porcine circoviruses, Asfarviridae and much more.