Microbial Physiology and Biochemistry Laboratory

• 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

'...Each experiment is set out with clearly defined goals, a brief introduction, and the materials that will be required...This is an adequate laboratory manual with a wide range of experiments...' David J.Kelly, Jnl of Antimicrobial Chemotherapy (1998) 42