The first libraries of complementary DNA (cDNA) clones were con structed in the mid-to-late 1970s using RNA-dependent DNA polymerase (reverse transcriptase) to convert poly A* mRNA into double-stranded cDNA suitable for insertion into prokaryotic vectors. Since then cDNA technology has become a fundamental tool for the molecular biologist and at the same time some very significant advances have occurred in the methods for con structing and screening cDNA libraries. It is not the aim of cDNA Library Protocols to give a comprehensive review of all cDNA library-based methodologies; instead we present a series of up-to-date protocols that together should give a good grounding of proce dures associated with the construction and use of cDNA libraries. In deciding what to include, we endeavored to combine up-to-date versions of some of the most widely used protocols with some very usefiil newer techniques. cDNA Library Protocols should therefore be especially useful to the investigator who is new to the use of cDNA libraries, but should also be of value to the more experienced worker. Chapters 1-5 concentrate on cDNA library construction and manipula tion, Chapters 6 and 7 describe means of cloning difficult-to-obtain ends of cDNAs, Chapters 8-18 give various approaches to the screening of cDNA libraries, and the remaining chapters present methods of analysis of cDNA clones including details of how to analyze cDNA sequence data and how to make use of the wealth of cDNA data emerging from the human genome project."

This thoughtful and provocative book provides a concise, up-to-date presentation of how current and projected future phosphorus scarcity will affect legume growth and their symbiotic nitrogen-fixing capabilities. It is a timely examination of the physiological and molecular responses of nodules to phosphorous deficiency in attempt to identify common principles. Students and researchers in the many disciplines related to crop productivity will find this title an exciting contribution in the area of plant stress physiology. The knowledge in this volume can also aid plant breeders, particularly through new methods of genetic engineering, in developing unique and adaptive cultivars with higher symbiotic efficiency. The awareness of the rapidly rising world population must translate into a parallel increase in agricultural production in order to sustain the growing population both now and in the future. Hence, the demand for food crops to produce proteins and vegetable oil for human consumption is going to increase considerably during the coming years. The essential role of legumes in agriculture is well-recognized, given the abundant levels of proteins and oils found in plants along with their enormous contribution to the sustainability of agricultural systems and human health. The capacity of legumes to fix nitrogen (N2) in partnership with rhizobia provides an input-saving and resource-conserving alternative, thereby reducing the need for chemical fertilizers while enhancing overall crop productivity. The use of N2-fixing legumes to produce plant proteins results in a substantial decrease in the consumption of fossil fuels and therefore also in the agricultural effects to global warming. However, a major constraint to legume production is low soil phosphorus (P) availability, considering that an overwhelming majority of the world's soils are classified as P-deficient. Low-P availability is especially problematic for legumes, since legume nodules responsible for N2 fixation have a high P requirement. Therefore, this book explains how nodule N2 fixation responds to low P availability, which is crucial for improving legume production and maintaining agricultural sustainability in the context of the global P crisis.

Over the past 10 years great progress has been made in the development of efficient techniques for both gene isolation and mapping. The identifica- tion and isolation of transcribed sequences from large chromosomal regions are central to the human genome mapping project. Techniques for isolating novel cDNAs have applications both in the overall construction and integra- tion of long-range physical and transcription maps and in the identification of disease genes. A number of different techniques for the isolation of cDNAs from mam- malian genomes have been developed, including screening "zoo" blots, the use of large genomic clones (YACs or cosmids) for hybridization against cDNA libraries, and CpG island mapping. More recently two highly efficient tech- niques have been introduced: exon trapping, based on the presence of exon splice sites, and direct selection, based on the enrichment of selected cDNAs using immobilized YACs or cosmids. Leading researchers in the field have contributed chapters detailing the practical procedures for these and other widely used methods. The most rapid progress presently being made in the field of gene isolation concerns the partial sequencing of cDNA clones from one or both ends to produce expressed sequence tags (ESTs). Indeed, by Octo- ber 1995, the EST division of Genbank (dbEST) contained a total of approxi- mately 270,000 human EST sequences accounting for almost half the number of sequence entries in Genbank.

In the past few years there has been the increased recognition that the effects of oxidative stress are not limited to the damage of cellular constituents. There is now evidence that reactive oxygen species (ROS) can alter cell function by acting upon the intermediates, or second messengers, in signal transductions. Such effects on signaling mechanisms probably account for the role of oxidative stress in inflammation, aging, and cancer. This volume brings together internationally recognized researchers in both the major areas covered by the book, oxidative stress and signal transduction. The work is organized in three sections. The first deals with the immediate cellular responses to oxidative stress and the production of second messengers. The second details the connection between second messengers and the gene. The third part looks more closely at the level of the gene.

This book features a special subsection of Nanomedicine, an application of nanotech nology to achieve breakthroughs in healthcare. It exploits the improved and often novel physical, chemical and biological properties of materials only existent at the nanometer scale. As a consequence of small scale, nanosystems in most cases are efficiently uptaken by cells and appear to act at the intracellular level. Nanotechnology has the potential to improve diagnosis, treatment and follow-up of diseases, and includes targeted drug delivery and regenerative medicine; it creates new tools and methods that impact sig nificantly upon existing conservative practices. This volume is a collection of authoritative reviews. In the introductory section we define the field (intracellular delivery). Then, the fundamental routes of nanode livery devices, cellular uptake, types of delivery devices, particularly in terms of localized cellular delivery, both for small drug molecules, macromolecular drugs and genes; at the academic and applied levels, are covered. The following section is dedicated to enhancing delivery via special targeting motifs followed by the introduction of different types of intracellular nanodelivery devices (e.g. a brief description of their chemistry) and ways of producing these different devices. Finally, we put special emphasis on particular disease states and on other biomedical applications, whilst diagnostic and sensing issues are also included. Intracellular delivery / therapy is a highly topical which will stir great inter est. Intracellular delivery enables much more efficient drug delivery since the impact (on different organelles and sites) is intracellular as the drug is not supplied externally within the blood stream. There is great potential for targeted delivery with improved localized delivery and efficacy.

Products from Cells - Cells as Products This book ist he "lasting" product, a resource ofup to date information in the scientific literature fort he field ofanimal cell tec hnology, as it was presented during a pleasant and s timulating mee ting that was held in Lugano Switzerland in April 1999. "Products" appeartwice int he title oft he conference. This clearly indicates the fact that the focus oft he papers presented during this meeting was really the application ofn ew technologies (novel reactors or novel vectors, for example for the preparation and/ort he more efficient generation ofproducts ) that could be used, mainly, int he medical field. Classical approaches forthe use ofa nimal cells, for example forthe p r oduction of virus vaccines for human and animal health, still remain an important technology and still have, surprisingly, quite significant potential for further development and improvement. How ever, it appears that major technological advances an d major growth from an economical point ofview are occurring in other areas. Most importantly, protein production on the basis of recombinant DNA molecules transferred into a nimal cells, appears to be an ever increasing field of interest and innovation, even though the first production scheme with this technology was approved more than 15 years ago.

The goal of Biological Aging: Methods and Protocols is to present some of the most promising and important tools that are currently used in biological aging research. These tools include established protocols such as aging cell culture as well as many more contemporary approaches such as nuclear transfer, microarray and proteomics technologies and the use of ribozymes in aging research. Collectively, these powerful tools combined with the many other techniques that are presented are rapidly advancing the exciting and expanding field of biological aging.

Despite many technological challenges faced by the xenotransplantation field, many major advances have been made in the last two decades. The field seeks to overcome the limitations and difficulties in organ procurement, which also apply to human cells and tissues, and facilitate the development of new therapies based on cell and engineered-tissue. Xenogeneic cells are simpler than solid organs and seem to pose less hurdles to attain long-term graft survival. In, Xenotransplantation: Methods and Protocols expert researchers study characterizations of xenogeneic interactions at the cellular and molecular levels and describe the use of relevant small-animal and pig-to-primate models. Related ethical and legal considerations are also covered. Written in the highly successful Methods in Molecular Biology (TM) series format, the chapters include the kind of detailed description and implementation advice that is crucial for getting optimal results in the laboratory. Thorough and intuitive, Xenotransplantation: Methods and Protocols aids scientists in continuing to study xenotransplantation and its multiple aspects.

Epithelia are one of the commonest tissue types in the animal kingdom. Chapters from leading scientists in the major international research laboratories use examples from different systems to illustrate the form and function of epithelia. An important theme is the way in which epithelial cells differentiate to specialized tissue - reversal of this process occurs when cells become tumorigenic.

Lysosomes are membrane-surrounded organelles which are present in all animal cells. The importance of this organelle is underlined by an increasing number of human diseases, which are associated with an impaired function of the lysosomal compartment.

This book summarizes the current state-of-the art knowledge about this unique organelle. It addresses the biogenesis of this compartment, the transport of lysosomal proteins, the role of the lysosomal membrane in lysosomal stability and transport, the function of lysosomal proteases and hydrolases, lysosomal storage disorders, and new concepts on how to treat these diseases. In addition to these classical topics, new insights into lysosomal functions are covered by chapters dealing with specialized lysosomes involved in bone resorption and plasma membrane repair, the lysosomal transciptome, and proteome and the emerging role of lysosomes in special forms of autophagy.

This book will provide readers with a comprehensive overview into how this fascinating organelle works and how research in the field is developing.

The plant' is often the most neglected part of plant-based medicine. Throughout time, humans have searched, collected, and effectively used plants for healing. Currently, the medicinal plant-based business is flourishing at a dramatic pace and at the expense of an already declining population of plant species, many of which are on the verge of extinction. In spite of this history and popularity, the mystery of what transforms a plant into a medicinal plant persists, and there are chronic problems with ensuring the safety and efficacy of medicinal plant products. Therefore, there is a real need for a full characterization of medicinal plant species and for the development and application of novel technologies for the production of plant-based medicines. This book highlights some of the recent advances and new approaches to the development of technologies for plant-based medicines and is intended to stimulate new discussions among researchers, regulatory authorities, and pharmaceutical organizations, leading to significant advancements in the field.

The rapidly developing field of nanomaterials has expanded in many commercial areas. More recent studies have begun to provide a foundation for understanding how nanomaterials influence cells and how they also can serve as methodological tools for studies in medicine and cell biology, including research into stem cells. Recent investigations have shown affects of nanomaterials on specific subcellular structures, such as the actin-based brush border network in cells with an increasing emphasis on the barrier function of epithelial tissues. While other studies have shown involvement of nanoparticles in specific cytoplasmic signal transduction events such as the rise in intracellular free calcium, a signaling event known to regulate many changes in cell architecture and function. In parallel, nanomaterials are increasingly used in medicine for drug delivery, treatment of cancer and an increasing number of new applications. This book investigates these areas and also includes new methods for assessment in cell biology and medicine.

This book focuses on issues in clinical practice and research that are of general interest. The articles primarily focus on understanding the pathogenic mechanisms of diseases, their prevention, and therapy. The topics addressed include cardiovascular regulation with regard to blood pressure and heart rate variability, and to coupling blood pressure changes with subarachnoid fluid oscillations. In addition, the book discusses recent advances in the diagnostics of and targeted molecular therapy for renal and pancreatic malignancies, growth disorders, vitamin D and calcium homeostasis in children in the context of neonatal urolithiasis, and neurosurgical interventions in multifarious age-related diseases of the vertebrae. Neuropsychological aspects of patients' quality of life and of shaping medical staff's attitude toward patients are also addressed. The respective articles are intended to build a bridge between basic and clinical research. Further, the book enhances the current body of knowledge on diagnostics and patient treatment and offers valuable new perspectives on practical clinical issues. As such, it offers a unique resource for clinicians, family physicians, medical scholars, and professionals engaged in patient management.

Recent work has revealed that stabilizing G-quadruplexes in telomeric DNA inhibits telomerase activity, providing impetus for the development of G-quartet-interacting drugs, while G-quartet-containing oligonucleotides have been recognized as a potent class of aptamers effective against STAT3 and other transcription factors implicated in oncogenesis, proving these guanine-quartets to be a vital and rich area for future study. In "G-Quadruplex DNA: Methods and Protocols", experts in the field present a collection of detailed techniques for studying G-quartet formation, dynamics, and molecular recognition. Written in the highly successful Methods in Molecular Biology (TM) series format, chapters include brief introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, "G-Quadruplex DNA: Methods and Protocols "promises to be a useful resource for those familiar with G-quartets as well as an easy entry point for those researchers from diverse fields who are just developing an interest in the exciting implications of G-quadruplex DNA.

The decline of infections, starvation, heart attack, and stroke has allowed people to reach extreme old age--and ushered in disability, dementia, and degenerative disease, with profound consequences for the self and society. In chapters echoing Dante's nine circles of hell, Dr. Guy Brown explores these vital issues at various levels, from the cell, to the whole body, to society and how all this new medical technology affects the meaning of death. He tracks the seismic shifts in the causes and character of death that are rocking medicine and reveals how technological innovations, such as cloning and electronic interfaces, hint at new modes of "survival" after death.

Nanoscale structures and materials have been explored in many biological applications because of their novel and impressive physical and chemical properties. Such properties allow remarkable opportunities to study and interact with complex biological processes. This book analyses the state of the art of piezoelectric nanomaterials and introduces their applications in the biomedical field. Despite their impressive potentials, piezoelectric materials have not yet received significant attention for bio-applications. This book shows that the exploitation of piezoelectric nanoparticles in nanomedicine is possible and realistic, and their impressive physical properties can be useful for several applications, ranging from sensors and transducers for the detection of biomolecules to sensible substrates for tissue engineering or cell stimulation. The book also focuses on the preparation, characterization and bio-applications of piezoelectric nanoparticles.

The current explosive progress in molecular biological research can be definitively traced to the development of molecular cloning technology. The ability to insert specific gene sequences into cloning vectors and their subse quent expansion is the cornerstone of modem molecular biology. A direct practical outcome of molecular cloning technology is its application to ex press specific recombinant genes. Currently, recombinant gene products are used in a wide spectrum of applications, including gene therapy, production of bioactive pharmaceuticals, synthesis of novel biopolymers, in agriculture and animal husbandry, and so on. A fundamental requirement for successful recombinant gene expression is the design of the cloning vector and the choice of the host organism for expression. Recombinant Gene Expression Protocols grows out of the need for a laboratory manual that provides the reader the background and rationale, as well as the practical protocols for the preparation of "expression constructs" and their introduction into appropriate host cells and/or organisms. The chap ters in this book are grouped by their expression hosts, including E. coli, yeast, mammalian cells, nonmammalian eukaryotes such as plants, Xenopus, and insects, as well as in transgenic organisms. In-depth information is presented on the important characteristics of expression cloning vectors and the various methods for efficiently introducing expression constructs into target cells and/ or organisms. Throughout Recombinant Gene Expression Protocols, the authors have consistently striven for a balanced presentation of both background informa tion and actual laboratory details.

Signalling and Communication: An Introduction to Section 1; R. Paton. Some Aspects of Gap Junction Dynamics in Embryonic Systems; S. Baigent, et al. Real Time Processing of Nerve Signals for Controlling a Limb Prostheses; M. Bodgan, W. Rosentiel. Stimulus-Secretion Coupling in Pancreatic beta-Cells Explained by Chay's Store-Operated Model; T.R. Chay. Towards Computational Models of Chemotaxis in Escherichia Coli; L. Clarke, R.C. Paton. Three Modes of Calcium-Induced Calcium Release (CICR) in Neurons; D.D. Friel. Theta-Neuron, a One Dimensional Spiking Model that Reproduces in Vitro and in Vivo Spiking Characteristics of Cortical Neurons; B.S. Gutkin, G.B. Ermentrout. Computation and Information: An Introduction to Section 2; R. Paton. Morphomechanical Feedback in Embryonic Development; L.V. Beloussov. Information Processing in Computational Tissues;M.H. Butler, et al. Semiotics of Complex Systems: A Hierarchical Notation for the Mathematical Structure of a Single Cell; J.L.R. Chandler. Localization and Nonlocality in Computation; R. Cottam, et al. Mining the Gene Expression Matrix: Inferring Gene Relationships from Large Scale Gene Expression Data; P. D'haeseleer, et al. Epilogue-Concluding Discussions; M. Holcombe. 18 Additional Chapters. Index.

This book outlines the interaction of cadmium with the proteome and signalling molecules of mammalian cells. Chapters from expert contributors cover topics such as cadmium chemical biology, membrane receptors and transporters for cadmium and cadmium complexes, and targets of cadmium toxicity. Students and researchers working in bioinorganic chemistry will find this book an important account.

This detailed volume provides thorough protocols describing how to use genetics to study mouse and zebrafish erythropoiesis in whole animal models and for genetically manipulating cultured mouse and human erythroid cells. Protocols include strategies to analyze circulating red blood cell parameters, as well as progenitor cell status, stage of differentiation, and enucleation. Methods for the study of erythroid cell gene regulation using chromatin immunoprecipitation and chromatin conformation capture are detailed. Furthermore, protocols are provided to examine stress erythropoiesis, erythroblastic islands, and erythroblast structure. The collection also includes reviews on the available mouse models of erythropoiesis, as well as on future considerations for good manufacturing practice in order to translate the manufacture of erythrocytes to the clinic. 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, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Erythropoiesis: Methods and Protocols provides a definitive source for molecular and cellular biology protocols to study erythropoiesis and to move this vital field forward.

The latest edition in this continuing series includes the newest advances in the rapidly evolving field of animal cell culture, genetic manipulations for heterologous gene expression, cell line enhancements, improved bioreactor designs and separations, gene therapy manufacturing, tissue engineering, anti-apoptosis strategies and cell cycle research. The contents include new research articles as well as critical reviews on emerging topics such as viral and viral-like agent contamination of animal cell culture components. These papers were carefully selected from contributions by leading academic and industrial experts in the biotechnology community at the recent Cell Culture Engineering VI Meeting in San Diego, USA, 1998. However, the book is not merely a proceedings. Audience: Biochemical engineers, cell biologists, biochemists, molecular biologists, immunologists and other disciplines related to cell culture engineering, working in the academic environment and the biotechnology or pharmaceutical industry.

This book describes the methods of analysis and determination of oxidants and oxidative stress in biological systems. Reviews and protocols on select methods of analysis of ROS, RNS, oxygen, redox status, and oxidative stress in biological systems are described in detail. It is an essential resource for both novices and experts in the field of oxidant and oxidative stress biology.

Histochemistry and cytochemistry are important fields for studying the inner workings of cells and tissues of the body. While visualization techniques have been in use for many years, new methods of detection developed recently help researchers and practitioners better understand cell activity. Histochemical and Cytochemical Methods of Visualization describes the essential techniques that can be used for histochemical investigations in both light and transmission electron microscopy. The book begins by discussing techniques in light microscopy. It reviews classical methods of visualization, histochemical and histoenzymatic methods, and methods used to visualize cell proliferation and apoptosis. Next, the book examines the cytochemical methods used in electron microscopy with traditional techniques, as well as more specialized methods. The final section provides an overview of image analysis and describes how image processing methods can be used to extract vital information. A 16-page insert supplies color illustrations to enhance the text. Techniques will continue to adapt to the latest technological innovations, allowing more and more precise quantification of images. These developments are essential to the biological as well as the medical sciences. This manual is a critical resource for novice and experienced researchers, technicians, and students who need to visualize what happens in the cell, the molecules expressed, the main enzymatic activities, and the repercussions of the molecular activities upon the structure of the cells in the body.

The "cancer stem cell" hypothesis postulates that cancer arises from a subpopulation of tumor-initiating cells or cancer stem cells (CSCs). While the idea of cancer stem cells has been around for more than a hundred years, evidence from the fields of hematology and cancer biology has now demonstrated the critical role of stem cells in hematological malignancies and suggested that these same mechanisms are also central to the initiation, progression, and treatment of solid cancers. Clinical and experimental studies have shown that CSCs exhibit many classical properties of normal stem cells, including a high self-renewal capacity and the ability to generate heterogeneous lineages; the requirement for a specific "niche"/microenvironment to grow; and an increased capacity for self-protection against harsh environments, toxins, and drugs. Cancer Stem Cells in Solid Tumors represents a detailed overview of cancer stem cells and their role in solid cancers. Comprised of 24 chapters, this volume will provide readers with a comprehensive understanding of this important and evolving field. Topics covered include: Introduction of the CSC hypothesis Historical perspectives and the contributing lessons from leukemia Current knowledge regarding the identification and role of CSCs in various forms of solid cancer including breast, brain, colorectal, pancreatic, prostate, melanoma, lung, ovarian, hepatocellular, and head and neck cancer Molecular pathways involved in driving CSC function, with a particular focus on the novel convergence of embryonic and tumorigenic signaling pathways In vitro and in vivo assays, model systems, and imaging modalities for studying CSCs The clinical importance of CSCs for cancer management and treatment, including important implications for prognosis, prediction, and treatment resistance Consideration of the controversy surrounding the CSC hypothesis and important unanswered questions in this field This collective work was written by a group of prominent international experts in cancer biology, oncology, and/or stem cell biology. It will serve as a valuable resource for established researchers, professors, health care professionals, and students in the medical and scientific community who are investigating stem cells and/or oncology.

he biological sciences are dominated by the idea that cells are the functionally autonomous, physically separated, discrete units of life. TThis concept was propounded in the 19th century by discoveries of the cellular structuring of both plants and animals. Moreover, the ap parent autonomy of unicellular eukaryotes, as well as the cellular basis of the mammalian brain (an organ whose anatomy for a long while defied attempts to validate the idea of the cellular nature of its neurons), seemed to provide the final conclusive evidence for the completeness of *cell theory', a theory which has persisted in an almost dogmatic form up to the present day. However, it is very obvious that there are numerous observations which indicate that it is not the cells which serve as the basic units of biological life but that this property falls to some other, subcellular assemblage. To deal with this intricate problem concerning the fundamental unit of living matter, we proposed the so-called Cell Body concept which, in fact, devel ops an exceedingly original idea proposed by Julius Sachs at the end of the 19th century. In the case of eukaryotic cells, DNA-enriched nuclei are intimately associated with a microtubular cytoskeleton. In this configuration-as a Cell Body-these two items comprise the fundamental functional and struc tural unit of eukaryotic living matter. The Cell Body seems to be inherent to all cells in all organisms.