Microinjection and Transgenesis

The technique of microinjection is routinely used to transfer biological materials - proteins, oligonucleotides, RNA and DNA - into cultured somatic cells and embryos. This manual provides over 100 protocols for microinjection and useful auxillary techniques, as well as the most relevant strategies that make this field a highly productive area of biological and biotechnological research. Special topics include: the use of YACs to produce transgenic mice, ligand-dependent site-specific recombination in embryonic stem cells, morula aggregation to generate germline chimeras as well as the analysis of injected cells.

The most potent technique to study cell function is the specific tranfer of genetic material into a single cell. Several functions can than be analyzed in the resulting transgenic cell or organism. The method of microinjection and transgenesis is currently used in many cell biological laboratories.

A practical tool presenting the latest techniques developed in renowned laboratories Easy to follow step-by-step protocols Background information and full practical details (materials needed, exact recipes, troubleshooting guide, typical results, list of suppliers, etc.)

Contenu
1 Strategies and Protocols for Microinjection Experiments Using Somatic Cells in Culture.- 1 Protocols for the Manual and Automatic Microinjection of Somatic Cells in Culture and for the Analysis of Microinjected Cells.- 2 Automated Computer-Assisted Microinjection into Cultured Somatic Cells.- 3 Immunofluorescence and Immunoelectron Microscopy of Microinjected and Transfected Cultured Cells.- 4 Videomicroscopy of Living Microinjected Hippocampal Neurons in Culture.- 5 Preparation, Purification, and Analysis of Synthetic Oligonucleotides Suitable for Microinjection Experiments.- 6 Colorimetric in Situ Hybridization.- 7 Manual Microinjection: Measuring the Inhibitory Effect of HIV-1-Directed Hammerhead Ribozymes in Tissue Culture Cells.- 8 Cellular Microbiochemistry.- 9 A New Strategy for Studying Microtubule Transport and Assembly During Axon Growth.- 10 The Use of Microinjection to Study Signal Transduction in Mammalian Cells.- 2 Strategies and Protocols for Microinjection Experiments Using Embryos.- 11 Supply and Husbandry of Mice for Transgenic Science.- 12 Mouse Male Germ Cells in Culture: Toward a New Approach in Transgenesis?.- 13 Sperm-Mediated Gene Transfer.- 14 Application of Ligand-Dependent Site-Specific Recombination in ES Cells.- 15 A Modified Protocol for the Generation of Transgenic Mice and Freezing of Mouse Embryos.- 16 The Use of Morula Aggregation to Generate Germline Chimeras from Genetically Modified Embryonic Stem Cells.- 17 The Generation of Transgenic Mice with Yeast Artificial Chromosomes.- 18 Following Up Microinjected mRNAs: Analysis of mRNA Poly(A) Tail Length During Mouse Oocyte Maturation.- 19 Mouse Preimplantation Embryos and Oocytes as an In Vivo System to Study Transcriptional Enhancers.- 20 The Use of Mouse Preimplantation Embryos for the Identification of DNA-Binding Proteins Essential in Early Development.- 21 Application of Transgenes and Transgenic Mice to Study Gene Activity from the Oocyte to the Early Embryo.- 22 Transitory Transgenic Analysis as an In Vivo System to Study Promoter Regulatory Elements.- 23 Transgenic Analysis of Embryonic Gene Expression Using LacZ as a Reporter.- 24 Autonomous Cell Labeling Using a laacZ Reporter Transgene to Produce Genetic Mosaics During Development.- 25 Multiple Developmental Functions of the Zinc Finger Gene Krox-20 Are Revealed by Targeted Insertion of the E. coli lacZ Coding Sequence.- 26 Transgenic Mouse Strategies in Virus Research.- 27 Isolation of Mutated Genes from Transgene Insertion Sites.- 28 Contributions of Transgenic and Knockout Mice to Immunological Knowledge.- 29 Generation and Application of Transgenic Rabbits.- 30 Microinjection of Drosophila Eggs.