Microbial Physiology and Biochemistry Laboratory A Quantitative Approach

About this book

This lab manual is intended as an adjunct to David White's Physiology and Biochemistry of Prokaryotes, for the laboratory component of a microbial physiology course. There is no other laboratory manual for teaching microbial physiology.

Contents

Introduction; Acknowledgements; Laboratory Rules and Safety; Experiment 1: An exercise in pipetting and the Beer-Lambert Law; Experiment 2: Bacterial growth curve; Experiment 3: Bioassay for micotinic acid; Experiment 4: The effect of environment on growth; Experiment 5: Lactic acid production by lactic acid bacteria; Experiment 6: Assay of protein and RNA in whole cells grown at different growth rates; Experiment 7: Analysis of diauxic growth; Experiment 8: Assay of amylase and protease secreted by Bacillus Subtilis; Experiment 9: Concentration of amylase from Bacillus Subtilis by ammonium sulfate precipitation and separation from protease by affinity purification; Experiment 10: Ion-exchange chromatography of amylase; Experiment 11: Induction of alkaline phosphatase and the determination of its cellular cation in Escherichia coli; Experiment 12: Assay of threonine deaminase: Determination of KM and Vmax; Experiment 13: Assaying fructose-1,6-bisphosphatase in Saccharomyces cerevisiae; Experiment 14: Purification of glucose-6-phosphate dehydrogenase; Experiment 15: How E. coli adapts to anaerobiosis: Nitrate reductase; Experiment 17: Chemotaxis of Pseudomonas aeruginosa; Experiment 18: Phototasix of Rhodospirllum centenum; Experiment 19: Light production by Phorobacterium phosphoreum; Appendix A: Analysis of experimental data: How to do simple dilutions; Guidelines for writing laboratory reports; Appendix B: Laboratory supplies and equipment; Appendix C: Media; Buffers; Solutions; Appendix D: RNA assay using orcinol; Lowry protein assay; Bradford Protein assay; Assay for *a-amylase using starch conjugated to a dye; Appendix E: The Beer-Lambert law; Relative absorbancies; Cell density vs. Klett units; Appendix F: Quantitative problems; Appendix G: Solutions to problems in Appendix F; Appendix H: Determining the amount of inorganic phosphate that limits the growth yields of E. coli, Factors that affect the relative amounts of saturated; unsaturated, and branched chain fatty acids; Measuring the KM and Vmax of fructose-1,6-bisphosphatase and the inhibition by AMP; Assaying glycolytic enzymes and fructose-1,6-bisphosphatase in yeast grown on glucose or ethanol; Regulation of phosphofructokinase activity; Further experiments and demonstrations with Photobacterium

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