MICR3100 lab manual .docx - MICR 3100/BIOL 3100 GENERAL MICROBIOLOGY LABORATORY EXERCISES Last Updated in Fall 2018 by Hyunsook Park Ph.D Department of

MICR3100 lab manual .docx - MICR 3100/BIOL 3100 GENERAL...

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Last Updated in Fall 2018 by Hyunsook Park, Ph.D. Department of Biological Sciences MICR 3100/BIOL 3100 GENERAL MICROBIOLOGY LABORATORY EXERCISES
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TABLE OF CONTENTS LABORATORY RULES ......................................................................................................................................... 3 LABORATORY SAFETY ................................................................................................................................... 3 HOUSE KEEPING ........................................................................................................................................... 4 EXERCISE 1. UBIQUITY OF MICROORGANISMS ............................................................................................... 6 EXERCISE 1-1. SAMPLING MICROBES IN THE ENVIRONMENT ..................................................................... 7 EXERCISE 1-2. OBSERVATION OF MICROBIAL COLONIES ............................................................................. 9 EXERCISE 2. TRANSFER AND ISOLATION OF MICROORGANISMS ................................................................. 12 EXERCISE 2-1. ASPETIC TECHNIQUE ........................................................................................................... 13 EXERCISE 2-2. 4-QUADRANT STREAKING ................................................................................................... 14 EXERCISE 2-3. MOTILITY AGAR INOCULATION (DEMO ONLY) ................................................................... 18 EXERCISE 3. MICROSCOPY ............................................................................................................................. 21 EXERCISE 4. STAINING MICROORGANISMS ................................................................................................... 26 EXERCISE 4-1. SIMPLES STAINING OF CORYNEBACTERIA SP ...................................................................... 28 EXERCISE 4-2. GRAM STAINING ................................................................................................................. 30 EXERCISE 5. EFFECTS OF THE ENVIRNMENTS ON MICROBIAL GROWTH ..................................................... 34 EXERCISE 5-1. TEMPERATURE AND MICROBIAL GROWTH ........................................................................ 35 EXERCISE 5-2. ATMOSPHERE - OXYGEN ..................................................................................................... 37 EXERCISE 5-3. pH AND MICROBIAL GROWTH ............................................................................................ 40 EXERCISE 5-4. OSMOLARITY AND MICROBIAL GROWTH ........................................................................... 41 EXERCISE 6. BASIC MICROPIPETTING PRACTICE ........................................................................................... 43 EXERCISE 7. BACTERIAL GROWTH CURVE ..................................................................................................... 47 EXERCISE 8. BACTERIAL TRANSFORMATION ................................................................................................. 55 EXERCISE 9. VIRAL INFECTION CYCLE ............................................................................................................ 61 EXERCISE 10. IDENTIFICATION OF UNKNOWN MICROORGANISMS ............................................................ 66 EXERCISE 10-1. ISOLATION OF GENOMIC DNA FROM UNKNOWN MICROORGANISMS ........................... 67 EXERCISE 10-2. AMPLIFICATION OF 16S rDNA CONSENSUS SEQUENCE BY POLYMERASE CHAIN REACTION (PCR) ......................................................................................................................................... 70 EXERCISE 10-3. PURIFICATION OF THE AMPLIFIED 16S rDNA FRAGMENT BY AGAROSE GEL ELECTROPHORESIS ..................................................................................................................................... 72 EXERCISE 10-4. BLASTANALYSIS TO IDENTIFY THE UNKNOWN ORGANISM .............................................. 75 1
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EXERCISE 11. ISOLATION OF ANTIBIOTIC PRODUCER FROM SOIL ................................................................ 77 EXERCISE 11-1. PRIMARY ISOLATION OF SOIL MICROORGANISMS ........................................................... 78 EXERCISE 11-2. ISOLATION OF PURE COLONIES ........................................................................................ 79 EXERCISE 11-3. TEST FOR ANTIBIOTIC PRODUCTION ................................................................................ 80 EXERCISE 12. THE FUNGI: YEASTS AND MOLDS ............................................................................................ 82 EXERCISE 12-1. IDENTIFYING FUNGI .......................................................................................................... 83 EXERCISE 12-2. YEAST DIFFERENTIATION .................................................................................................. 85 EXERCISE 13. WATER CONTAMINATION AND WATER QUALITY TEST ........................................................... 87 EXERCISE 14. BLOOD LEUKOCYTE DIFFERENTIATION ................................................................................... 91 EXERCISE 15. ANTISEPTICS AND ANTIMICROBIALS ...................................................................................... 95 EXERCISE 15-1. EFFECTIVENESS OF ANTISEPTICS ...................................................................................... 97 EXERCISE 15-2. ANTIBIOTIC SUSCEPTIBILITY TESTING: THE KIRBY-BAUER METHOD ................................ 99 EXERCISE 16. MICROBIOME: VIDEO REVIEW PRESENTATION .................................................................... 103 2
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LABORATORY SAFETY Lab coats must be worn at all times. Long hair must be tied back at all times- everyone! Closed toe shoes must be worn at all times- to protect from spills. No eating, chewing, drinking or smoking. Wear protective gloves when working with microbial cultures Wash hands with antimicrobial soap before you start and before you leave. Wash down bench with disinfectant before and after work. Only lab manual, no other extraneous books, materials should be on bench. Store these items under the bench or on shelves along the wall. Please report all spills and accidents immediately. DO NOT TRY TO CLEAN UP by yourself. Do not leave cultures or other experimental vials in drawers and cabinets unless told to do so by the instructor.
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