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OverviewUsing an integrated series of laboratory exercises that involve the cloning and analysis of the bioluminescence (lux) genes from the marine bacterium Vibrio fischeri, this manual introduces students to the basic techniques of modern molecular biology. By organizing the exercises as part of a major cloning project, students get the sense of performing a complete project, rather than just learning a collection of procedures. Full Product DetailsAuthor: Michael Winfrey , Marc Rott , Alan Wortman , Alan Wortman (all of University of Wisconsin, La Crosse, USA)Publisher: Pearson Education (US) Imprint: Pearson Dimensions: Width: 10.00cm , Height: 10.00cm , Length: 10.00cm Weight: 0.100kg ISBN: 9780132700344ISBN 10: 0132700344 Pages: 400 Publication Date: 10 February 1997 Audience: Professional and scholarly , Professional & Vocational Format: Paperback Publisher's Status: Active Availability: In Print  This item will be ordered in for you from one of our suppliers. Upon receipt, we will promptly dispatch it out to you. For in store availability, please contact us. Table of ContentsI. INTRODUCTORY TECHNIQUES. 1. Introduction to the Laboratory: Basic Equipment and Bacteriological Techniques. 2. Preparation of Media and Reagents Used in Molecular Biology. 3. Isolation of Luminescent Bacteria from Natural Sources. 4. Restriction Digestion and Agarose Gel Electrophoresis of DNA. II. DNA ISOLATION AND ANALYSIS. 5. Isolation of Chromosomal DNA from Vibrio fischeri. 6. Large-Scale Purification of Plasmid DNA. 7. Spectrophotometric Analysis of DNA. III. CLONING THE LUX OPERON 8. Restriction Digestion of Vibrio fischeri Genomic DNA and Plasmid Vector. 9. Quantification of Genomic DNA by Fluorometry and Agarose Plate Fluorescence. 10. Ligation of Restriction Fragments of Vibrio fischeri DNA to Plasmid Vector. 11. Preparation of Competent Escherichia coli DH5. 12. Transformation of Competent Escherichia coli DH5 with Recombinant Plasmids. 13. Screening the Vibrio fischeri Genomic Library for Light Producing Clones. IV. RESTRICTION MAPPING AND SOUTHERN BLOTTING 14. Small-Scale Plasmid Isolations (Mini-preps) from Bioluminescent Clones. 15. Restriction Mapping of Plasmids from Bioluminescent Clones. 16. Southern Blotting and Hybridization to Detect the luxA Gene. V. SUBCLONING THE LUX A GENE 17. Restriction Digestion of lux Plasmids and Cloning Vector for Subcloning luxA. 18. Gel Purification of DNA Restriction Fragments Containing luxA. 19. Subcloning luxA into A Plasmid Vector. 20. Transformation of Competent Escherichia coli DH5 with Subcloned DNA. 21. Colony Hybridization to Screen for luxA Subclones. 22. Small-Scale Plasmid Isolations (Mini-Preps) from luxA Clones. VI. ADVANCED TECHNIQUES. 23. Amplification of luxA from Natural Isolates by the Polymerase Chain Reaction (PCR). 24. Southern Blotting and Hybridization of PCR Products. 25. DNA Sequencing of lux Genes from Plasmid Templates. 26. Computer Analysis of DNA Sequences Using the World Wide Web. 27. Mapping the Vibrio fischeri Genome by Pulsed Field Gel Electrophoresis. 28. Independent Projects in Molecular Biology. APPENDICES. 1. The Metric System and Units of Measure. 2. Centrifugation. 3. Spectrophotometry. 4. Agarose and Polyacrylamide Gel Electrophoresis. 5. Methylene Blue Staining of Agarose Gels. 6. Nucleic Acid Hybridization. 7. Alcohol Precipitation of Nucleic Acids. 8. Care and Handling of Enzymes. 9. Restriction Endonucleases. 10. Enzymes Used in Molecular Biology. 11. Maps of Cloning Vectors and Bacteriophage Lambda. 12. Proper Handling and Disposal of Hazardous Materials. 13. Procedures and Precautions for the Use of Radioisotopes. 14. Equipment and Supplies. 15. Media and Reagents. 16. Bacterial Strains. 17. Lists of Suppliers. 18. Recommended References. 19. Restriction Mapping Problems.ReviewsAuthor InformationTab Content 6Author Website:Countries AvailableAll regions |
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