cDNA Library Protocols

cDNA Library Protocols
Author: Ian G. Cowell
Publisher: Springer Science & Business Media
Total Pages: 324
Release: 2008-02-02
Genre: Science
ISBN: 1592595553

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.

Yeast Protocols

Yeast Protocols
Author: Wei Xiao
Publisher: Springer Science & Business Media
Total Pages: 389
Release: 2008-02-03
Genre: Science
ISBN: 1592599583

In this second edition of a widely used classic laboratory manual, leading experts utilize the tremendous progress and technological advances that have occurred to create a completely new collection of not only the major basic techniques, but also advanced protocols for yeast research and for using yeast as a host to study genes from other organisms. The authors provide detailed methods for the isolation of subcellular components-including organelles and macromolecules, for the basic cellular and molecular analysis specific for yeast cells, and for the creation of conditional mutant phenotypes that lend themselves to powerful genome manipulation. Additional protocols offer advanced approaches to study genetic interactions, DNA and chromatin metabolism, gene expression, as well as the foreign genes and gene products in yeast cells.

Generation of cDNA Libraries

Generation of cDNA Libraries
Author: Shao-Yao Ying
Publisher: Springer Science & Business Media
Total Pages: 329
Release: 2008-02-03
Genre: Science
ISBN: 1592593593

Since its invention and subsequent development nearly 20 years ago, po- merase chain reaction (PCR) has been extensively utilized to identify numerous gene probes in vitro and in vivo. However, attempts to generate complete and full-length complementary cDNA libraries were, for the most part, fruitless and remained elusive until the last decade, when simple and rapid methods were developed. With current decoding and potential application of human genome information to genechips, there are urgent needs for identification of functional significance of these decoded gene sequences. Inherent in bringing these app- cations to fruition is the need to generate a complete and full-length cDNA library for potential functional assays of specific gene sequences. Generation of cDNA Libraries: Methods and Protocols serves as a laboratory manual on the evolution of generation of cDNA libraries, covering both ba- ground information and step-by-step practical laboratory recipes for which p- tocols, reagents, operational tips, instrumentation, and other requirements are detailed. The first chapter of the book is an overview of the basics of generating cDNA libraries, which include the following: (a) the definition of a cDNA library, (b) different kinds of cDNA libraries, (c) differences between methods for cDNA library generation using conventional approaches and novel stra- gies, including reverse generation of RNA repertoires from cDNA libraries, and (d) the quality of cDNA libraries.

Malaria Methods and Protocols

Malaria Methods and Protocols
Author: Denise L. Doolan
Publisher: Springer Science & Business Media
Total Pages: 606
Release: 2008-02-02
Genre: Medical
ISBN: 1592592716

The Plasmodium spp. parasite was identified as the causative agent of malaria in 1880, and the mosquito was identified as the vector in 1897. Despite subsequent efforts focused on the epidemiology, cell biology, immunology, molecular biology, and clinical manifestations of malaria and the Plasmodium parasite, there is still no licensed vaccine for the prevention of malaria. Physical barriers (bed nets, window screens) and chemical prevention methods (insecticides and mosquito repellents) intended to interfere with the transmission of the disease are not highly effective, and the profile of resistance of the parasite to chemoprophylactic and chemotherapeutic agents is increasing. The dawn of the new millennium has seen a resurgence of interest in the disease by government and philanthropic organizations, but we are still faced with compl- ities of the parasite, the host, and the vector, and the interactions among them. Malaria Methods and Protocols offers a comprehensive collection of protocols describing conventional and state-of-the-art techniques for the study of malaria, as well as associated theory and potential problems, written by experts in the field. The major themes reflected here include assessing the risk of infection and severity of disease, laboratory models, diagnosis and typing, molecular biology techniques, immunological techniques, cell biology techniques, and field applications.

Combinatorial Peptide Library Protocols

Combinatorial Peptide Library Protocols
Author: Shmuel Cabilly
Publisher: Springer Science & Business Media
Total Pages: 320
Release: 2008-02-02
Genre: Science
ISBN: 1592595715

During the course of evolution, an imbalance was created between the rate of vertebrate genetic adaptation and that of the lower forms of living organisms, such as bacteria and viruses. This imbalance has given the latter the advantage of generating, relatively quickly, molecules with unexpected structures and features that carry a threat to vertebrates. To compensate for their weakness, vertebrates have accelerated their own evolutionary processes, not at the level of whole organism, but in specialized cells containing the genes that code for antibody molecules or for T-cell receptors. That is, when an immediate requirement for molecules capable of specific interactions arose, nature has preferred to speed up the mode of Darwinian evolution in pref- ence to any other approach (such as the use of X-ray diffraction studies and computergraphic analysis). Recently, Darwinian rules have been adapted for test tube research, and the concept of selecting molecules having particular characteristics from r- dom pools has been realized in the form of various chemical and biological combinatorial libraries. While working with these libraries, we noticed the interesting fact that when combinatorial libraries of oligopeptides were allowed to interact with different selector proteins, only the actual binding sites of these proteins showed binding properties, whereas the rest of the p- tein surface seemed "inert. " This seemingly common feature of protein- having no extra potential binding sites--was probably selected during evolution in order to minimize nonspecific interactions with the surrounding milieu.

Methods in Genomic Neuroscience

Methods in Genomic Neuroscience
Author: Hemin R. Chin
Publisher: CRC Press
Total Pages: 414
Release: 2001-09-26
Genre: Science
ISBN: 100061171X

The past few years have witnessed extraordinary advances in molecular genetic techniques and the accumulation of structural genomics information and resources in both human and model organisms. With the development of new technologies and the availability of resources like the sequence of eukaryotic genomes, problems of a previously unthinkable sco

Natural Killer Cell Protocols

Natural Killer Cell Protocols
Author: Kerry S. Campbell
Publisher: Springer Science & Business Media
Total Pages: 394
Release: 2008-02-03
Genre: Medical
ISBN: 1592590446

In Natural Killer Cell Protocols: Cellular and Molecular Methods, Kerry S. Campbell and Marco Colonna have assembled a comprehensive collection of readily reproducible methods designed to study natural killer (NK) cells from the broadest variety of viewpoints. These include not only classic techniques, but also new approaches to standard methods, newly evolved techniques that have become valuable for specific applications, and unique models for manipulating and studying NK cells. Among the advanced methods covered are those for in vitro transendothelial migration, in vivo detection of cells migrating into tumors, immunofluorescence staining of intracellular cytokines, and in vitro NK cell development. Valuable techniques for specific applications include vaccinia virus protein expression, soluble KIR-Fc fusions for HLA class I binding assays, calcium mobilization in cell conjugates, and identification of heterodimeric receptor complexes using cDNA library expression cloning. No less important are accounts of such classic methods as hybrid resistance, ADCC, viral defense, target cell cytotoxicity assays, cloning and culturing, tumor immunotherapy, and generation of HLA class I transfected target cells. Natural Killer Cell Protocols: Cellular and Molecular Methods offers immunologists, cancer researchers, virologists, and cell biologists today's most comprehensive collection of both established and cutting-edge techniques, methods that will contribute significantly to advancing our understanding of this fascinating and critically important class of cells.

Novel Methods in Molecular and Cellular Biochemistry of Muscle

Novel Methods in Molecular and Cellular Biochemistry of Muscle
Author: Grant N. Pierce
Publisher: Springer Science & Business Media
Total Pages: 234
Release: 2012-12-06
Genre: Science
ISBN: 1461563534

Experimental techniques are the life blood of science. The better the methodology is, the more reliable and accurate the results will be. Ultimately, this will lead to a clearer interpretation of those results and firmer conclusions from any set of experiments. Experimental methodology in the area of cardiovascular biochemistry and molecular biology has advanced considerably in the last decade. Because of these factors, it was thought that a focused issue of Molecular and Cellular Biochemistry dedicated to the novel, latest technological advances in the field was warranted. We must thank Dr Naranjan S. Dhalla, Editor-in-Chief of Molecular and Cellular Biochemistry, for his willingness to publish an issue with such a focus. We have attracted some of the leaders in the field of cardiovascular biology to submit articles describing some of the most novel, significant techniques currently in use in their laboratories. The purpose of the manuscripts was not to describe the recent experimental findings from each laboratory as is done in most conventional manuscripts. Instead, the purpose of the articles found within this focused volume of Molecular and Cellular Biochemistry was to describe how the technique is performed on the laboratory bench so that others less familiar with the technique may be able to use it in their own labs. The subjects described in this volume can be generally subdivided into three categories: molecular biology, cell biology and basic biochemistry. The methods cover wide areas including various DNA and RNA expression technologies, transfection techniques, quantification of ion flux movement, measurements of lipid metabolism, advances in the culture of specific cardiovascular cell populations, and the use of confocal microscopy to examine cell structure and function. We thank all of the authors who have contributed so much of their time and efforts and, most importantly, shared the `secrets' of these valuable techniques with the rest of the cardiovascular research community.

Surgical Research

Surgical Research
Author: Wiley W. Souba
Publisher: Academic Press
Total Pages: 1526
Release: 2001-01-25
Genre: Health & Fitness
ISBN: 9780126553307

Contributors. -- Foreword. -- Preface. -- Getting Started. -- Assessing Available Information. -- Organizing and Preliminary Planning for Surgical Research -- Writing a Protocol: Animals, Humans, and Use of Biologic, Chemical, and Radiologic Agents. -- Grantsmanship. -- Informed Consent and the Protection of Human Research Subjects: Historical Perspectives and Guide to Current United States Regulations. -- Animal Care and Maintenance. -- Funding Strategies and Agencies: Academic-Industrial Relationships; Intellectual Property. -- Statistical Considerations. -- Use of Nonexperimental Studies to Evaluate Surgical Procedures and Other Interventions: The Challenge of Risk Adjustment. -- Measuring Surgical Outcomes. -- Design of Clinical Trials. -- Using Administrative Data for Clinical Research. -- Research in the Intensive Care Unit: Ethical and Methodological Issues. -- Research in the Operating Room. -- Effects of Age and Gender. -- Strategies, Principles, and Techniques Using Transgeni ...

Handbook of Molecular and Cellular Methods in Biology and Medicine, Second Edition

Handbook of Molecular and Cellular Methods in Biology and Medicine, Second Edition
Author: Leland J. Cseke
Publisher: CRC Press
Total Pages: 604
Release: 2003-11-24
Genre: Science
ISBN: 9780849308154

Since the publication of the best-selling Handbook of Molecular and Cellular Methods in Biology and Medicine, the field of biology has experienced several milestones. Genome sequencing of higher eukaryotes has progressed at an unprecedented speed. Starting with baker's yeast (Saccharomyces cerevisiae), organisms sequenced now include human (Homo sapiens), model crucifer (Arabidopsis thaliana), and rice (Oryza sativa). The invention of DNA microarray technology and advances in bioinformatics have generated vast amounts of genomic data. Reflecting these revolutionary advances Handbook of Molecular and Cellular Methods in Biology and Medicine, Second Edition documents conventional and modern approaches to tackle scientific research in the post-genomics era. Maintaining the step-by-step format that popularized the first edition, each chapter provides the principles behind the featured method, a detailed description of each protocol, applications of the protocol to different systems, and references for further study. Handbook of Molecular and Cellular Methods in Biology and Medicine, Second Edition now includes: New protocols in all chapters, including alternative protocols In vitro transcription methods Analysis of DNA sequences New bioseparation techniques New chapters covering: mRNA differential display Inhibition of gene expression In situ hybridization (Localization of gene expression) Combinatorial techniques Computational data mining methods applied to combinatorial chemistry libraries With this book at hand, researchers, teachers, and students can understand and utilize the major techniques and methods currently employed in cellular and molecular biology.