Guide to Electroporation and Electrofusion

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  • Format: Hardcover
  • Copyright: 1991-12-01
  • Publisher: Academic Pr

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Electroporation is an efficient method to introduce macromolecules such as DNA into a wide variety of cells. Electrofusion results in the fusion of cells and can be used to produce genetic hybrids or hybridoma cells. Guide to Electroporation and Electrofusion is designed to serve the needs of students, experienced researchers, and newcomers to the field. It is a comprehensive manual that presents, in one source, up-to-date, easy-to-follow protocols necessary for efficient electroporation and electrofusion of bacteria, yeast, and plant and animal cells, as well as background information to help users optimize their results through comprehension of the principles behind these techniques. Key Features * Covers fundamentals of electroporation and electrofusion in detail * Molecular events * Mechanisms * Kinetics * Gives extensive practical information * The latest applications * Controlling parameters to maximize efficiency * Available instrumentation * Presents applications of electroporation and electrofusion in current research situations * State-of-the-art modifications to electrical pulses and generators * Application of electroporation and electrofusion to unique, alternative cell and tissue types * Gives straightforward, detailed, easy-to-follow protocols for * Formation of human hybridomas * Introduction of genetic material into plant cells and pollen * Transfection of mammalian cells * Transformation of bacteria, plants, and yeast * Production of altered embryos * Optimization of electroporation by using reporter genes * Comprehensive and up-to-date * Convenient bench-top format * Approximately 125 illustrations complement the text * Complete references with article titles * Written by leading authorities in electroporation and electrofusion

Table of Contents

Prefacep. ix
Overview of Electroporation and Electrofusionp. 1
Mechanisms and Fundamental Processes in Electroporation and Electrofusion
Structure and Dynamics of Electric Field-Induced Membrane Pores as Revealed by Rapid-Freezing Electron Microscopyp. 9
Events of Membrane Electroporation Visualized on a Time Scale from Microsecond to Secondsp. 29
Time Sequence of Molecular Events in Electroporationp. 47
Electropores in Lipid Bilayers and Cell Membranesp. 63
Biophysical Considerations of Membrane Electroporationp. 77
Progress toward a Theoretical Model for Electroporation Mechanism: Membrane Electrical Behavior and Molecular Transportp. 91
Mechanisms of Electroporation and Electrofusionp. 119
Interfacial Membrane Alteration Associated with Electropermeabilization and Electrofusionp. 139
Membrane Fusion Kineticsp. 155
Effects of Intercellular Forces on Electrofusionp. 167
Dynamics of Cytoskeletal Reorganization in CV-1 Cells during Electrofusionp. 179
Applications of Electroporation and Electrofusion in Current Research
Gene Transfer into Adherent Cells Growing on Microbeadsp. 201
Gene Targeting and Electroporationp. 209
Pollen Electrotransformation for Gene Transfer in Plantsp. 227
Electrofusion of Plant Protoplasts and the Production of Somatic Hybridsp. 249
Electrotransformation of Bacteria by Plasmid DNAp. 265
Creating Vast Peptide Expression Libraries: Electroporation as a Tool to Construct Plasmid Libraries of Greater than 109 Recombinantsp. 291
Electroporation and Electrofusion Using a Pulsed Radio-Frequency Electric Fieldp. 303
Electroinsertion: An Electrical Method for Protein Implantation into Cell Membranesp. 327
Electroporation as a Tool to Study Enzyme Activities in Situp. 347
Comparison of PEG-Induced and Electric Field-Mediated Cell Fusion in the Generation of Monoclonal Antibodies against a Variety of Soluble and Cellular Antigensp. 363
Production of Genetically Identical Embryos by Electrofusionp. 371
Development of Cell-Tissue Electrofusion for Biological Applicationsp. 393
Novel Applications of Electroporationp. 411
Practical Protocols for Electroporation and Electrofusion
Design of Protocols for Electroporation and Electrofusion: Selection of Electrical Parametersp. 429
Protocols for Using Electroporation to Stably or Transiently Transfect Mammalian Cellsp. 457
Optimization of Electroporation Using Reporter Genesp. 465
Genetic Manipulation of Plant Cells by Means of Electroporation and Electrofusionp. 471
Protocols for the Transformation of Bacteria by Electroporationp. 485
Protocol for High-Efficiency Yeast Transformationp. 501
Protocols of Electroporation and Electrofusion for Producing Human Hybridomasp. 507
Human Hybridoma Formation by Hypo-Osmolar Electrofusionp. 523
Electrically Induced Fusion and Activation in Nuclear Transplant Embryosp. 535
Instrumentation for Electroporation and Electrofusion
Pulse Generators for Electrofusion and Electroporationp. 555
Indexp. 571
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