Microbiology Ambassadors High School Lab Experiments   Fall 1997
 

Contents Page

Magic Bullet Factories: Isolating Antibiotic Producing Bacteria
Abstract, Introduction, Special Materials, Time required 3
Preparation, Making the agar medium 4
Part I: Sampling the Soil or Compost 5
Part II: Selecting a Bacterium with a Zone of Clearing 5
Part III: Adding the Cross-Streaks 6
Part IV: Finding Antibiotic Producing Bacteria 6
Healing from the Kitchen
Abstract, Introduction, Special Materials, Time required 7
Preparation, Making the Agar Medium 8
Part I: Making the Extracts 9
Part II: A Drop of Healing 9
Part III: Checking for Growth Inhibitors 10
Do Soaps Really Do It?
Abstract, Introduction, Special Materials, Time required 11
Prep:  Making up the Plate Count Agar Medium 12
Part I: Asking a Question 12
Part II: Checking Results and Redesigning the Experiment 13
Genetic Exchange in Bacteria
Abstract, Introduction, Special Materials, Time required 14
Preparation: Making solutions and growing cultures 15
Part I: Plasmid Preparation 16
Part II: Making competent E.coli cells 17
Part III: the Transformation (genetic exchange) 17
Appendix, Culture Media, Supply Houses 19
 

"Magic Bullet" Factories: Isolating Antibiotic Producing Bacteria

Authors
Susan M. Merkel, Scott Kachlany, Kimberly Baldwin, Malik Williams
Section of Microbiology, Cornell University

Abstract:
This activity demonstrates where anti-microbial compounds come from by allowing the students to isolate antibiotic producing bacteria from soil or compost.  Students spread soil or compost onto agar plates, and incubate 1-2 days.   Students then test colonies that show a clearing zone against known Gram-positive and Gram-negative bacteria.  This lab activity was designed for and tested in middle or high school biology classrooms.

Introduction
Since the early 20th century, antibiotic compounds have been used across the world to fight against all kinds of bacterial diseases.  These low- molecular weight compounds are produced by microorganisms, such as fungi or bacteria, to kill off other bacteria that are competing for similar resources.  In general, antibiotics work by inhibiting specific bacterial enzymes.  In this experiment, students will obtain soil or compost from various sites to find naturally occurring antibiotic producing bacteria.

Special Materials:
 Soil or compost samples
 Petri plates with Plate Count or Starch -Casein Agar
 Some of the following bacteria (available from Ward's Biology:  1-(800) 962-2660 or Carolina Biological Supply:
 1-(800) 334-5551)
  Gram-negative  Gram-positive
  Escherichia coli  Staphylococcus epidermidis
  Pseudomonas sp.            Bacillus subtilis

Time Required:

Prep-Day Obtain soil or compost from various sites. Prepare agar petri plates.
 (Time: 1- 2 hours).

Day 1 Mix soil or compost with tap water in a clean bottle and shake.  Dip a sterile cotton swab into soil suspension.
Spread on agar and incubate overnight (Time: 10 minutes).

Day 2 Look at plate for zones of clearing and select those colonies. Pick a colony and streak a single line down the
center of the plate and incubate overnight
 (Time: 10- 15 minutes).

Day 3 With a toothpick, pick known test bacteria (Staphylococcus, E. coli, Pseudomonas, or Bacillus).  Streak bacteria
from the inner streak to the outer edges and incubate overnight (Time: 10- 15 minutes).

Day 4 Observe plates for any zone of inhibition near the center streak
 (Time: 10-15 minutes).

______________________________________________________________________________

Preparation:  Making the Agar Medium
Instructor or students need to make Plate Count Agar or Starch-Casien plates before isolating bacteria.  Alternatively, these items can be purchased from Ward's Biology: 1-(800) 962-2660 or Carolina Biological Supply: 1-(800) 334-5551)
NOTE:  This lab is written using Plate Count Agar.  Starch-Casein Agar produces more antibiotic producing isolates,
but they take 3-5 days to grow.  The recipe for Starch-Casein Agar is given in Appendix A.

Materials
The following recipe makes about 20 plates:
Equipment: 1 liter Erlenmeyer flask
  balance, weighing boats and spoons
  aluminum foil
  hot plate or autoclave
  protective gloves
  20 sterile petri plates
Supplies: 11.75 g of Plate Count Agar (or Starch-Casein agar)
  tap water

Procedure
1) Weigh out 11.75 g of Plate Count Agar (PCA) and place in Erlenmeyer flask.

2) Add 500 ml of distilled water to the flask and mix thoroughly.  Cover with aluminum foil.

3) Boil contents on hot plate for 30 minutes, or autoclave at 121oC for 20 minutes.   Let cool for 5 minutes or until
easy to handle.

4) With protective gloves, pour PCA mixture into 20 petri plates and let cool until solid (about 10 minutes)
NOTE:  Use caution when pouring plates.  To prevent contamination, only open the petri plate as long as
absolutely necessary.

5) Label plate "PCA".  Allow the plates to dry out by placing them at room temperature for 48 hours.
 

Part I: Sampling the Soil or Compost

Materials
Each group should be provided with the following:
Equipment:  2 clean bottles with lids
Supplies:  2 Plate Count Agar petri plate per soil sample
   2 sterile cotton swabs
   100 ml water

Procedure for Day 1
Experiment
1) Mix 1 tablespoon of soil or compost and 50 ml of tap water in a clean bottle and shake vigorously for 1 minute.
NOTE:   Be sure to dispose of all contaminated objects to a proper container.  To prevent contamination, only open
the petri plate as long as absolutely necessary.

2) Take a sterile cotton swab and dip into soil suspension.

3) With swab, spread out suspension onto PCA media.  To spread, pick a spot near the edge of the petri dish and with continuous, overlapping gentle strokes, cover the entire plate.

Control
4) Add 50 ml water to a clean bottle (the same water used to make the soils solution).

5) Take a sterile cotton swab and dip into water.

6) With swab, spread out the water onto PCA media (as described above).

7) Be sure to label each plate with your name (initials), date, and material that is being spread onto the dish.

8) Incubate plates overnight at room temperature.

______________________________________________________________________________

Part II: Selecting a Bacterium with a Zone of Clearing

Materials
Each group should be provided with the following:
Supplies: plates from the previous day
  8 sterile cotton swabs
  toothpicks
  8 PCA (Plate Count Agar) plates

Procedure for Day 2
1) Inspect plates for bacterial colonies surrounded by zones of clearing (see diagram).  This implies inhibition of growth
of other neighboring bacteria.

2) Use a toothpick to pick a colony at the center of the zone of inhibition, then gently streak the toothpick in a single line
down the center of a new PCA plate (Figure 3).

3) Incubate the plates overnight.

_______________________________________________________________________________

Part III: Adding the Cross Streaks

Materials
Each group should be provided with the following:
Supplies: toothpicks
  different strains of known test bacteria growing on PCA agar
  (we used Esherichia coli, Staph. epidermidis, Ps. aeruginosa  and Bacillus subtilis)

Procedure for Day 3
1) With a sterile toothpick, pick a colony from one of the known test bacteria plates.  Begin streaking the petri plate from the inner edges of the streak toward the outer edge of the plate (see figure), barely touching the initial streak made on Day 2.
NOTE:  Be careful when handling these and any bacteria.  Although these strains are not pathogenic, care should be taken to not spread them around.

2) On the same dish, repeat step 2 with the other known test bacteria.  Streak each bacterium about 1 inch from other streaks.  Label each known test bacterium.

3) Incubate the plates overnight at room temperature.

_______________________________________________________________________________

Part IV: Finding Antibiotic Producing Bacteria

Materials
Each group should be provided with the following:
Supplies: plates from Day 3

Procedure for Day 4
1) Observe plates for any inhibition of the known test bacteria.  A positive result is indicated by a lack of growth near the centerline.  (Why?)
 
 
 

Healing From the Kitchen

Authors:
Susan M. Merkel, Scott Kachlany, Kimberly Baldwin, Malik Williams
Section of Microbiology, Cornell University

Abstract:
This activity demonstrates how some researchers find new anti-microbial compounds by allowing the students to screen different herbs, spices, and plants for the ability to inhibit bacterial growth.   Student spread different bacteria onto agar plates, and test extracts from these various materials for growth inhibition using a Disk Diffusion Assay.  This lab activity was designed for and tested in middle or high school biology classrooms.

Introduction:
For more than 5,000 years, herbal remedies have been used in many different cultures to cure and inhibit disease.  People have claimed to successfully treat many different diseases, ranging from minor infections to cancer, with different herbal concoctions.  With the rise in antibiotic resistant bacteria, doctors need more antibiotic and growth inhibiting compounds.  Many researchers are now looking to natural products.  This activity shows students how natural products are screened for anti-bacterial activity.

Special Materials:
Equipment: weighing balance

Supplies: rubbing alcohol (or pure ethanol)
 small tubes with tight lids (eppendorf tubes work well)
 tetracycline antibiotic disks (available from
  Fisher Scientific Company 1-(800) 766-7000)
 1/4" sterile disks
 Petri plates with Plate Count Agar
 Gram(-) and Gram (+) bacteria types
  (We used Escherichia coli and Staphylococcus epidermidis, available from Ward's Biology: 1-(800) 962-2660
or
 Carolina Biological Supply: 1-(800) 334-5551)
 Some of the following plant materials (or anything else students want to test): teas; spices; herbs; plant leaves,
 stems or roots; mushrooms; etc. (NOTE: we got positive results with garlic, cloves and cumin)
 
Time Required
Prep Day Prepare the petri plates with Plate Count Agar.
 
Day 1 Weigh out material and let it sit in a known volume of alcohol overnight.

Day 2 Add one drop of each extract onto a sterile disk and allow to dry.  With sterile cotton swab, pick and spread
a known test bacterium onto Plate Count Agar.  Place dry disks on bacterial lawn and incubate overnight.

Day 3 Observe plates for any zones of clearing around the disk.  Clearing (growth inhibition) around the disk is
a positive result.
 

Prep:  Making the Agar Medium
Instructor or students need to make Plate Count Agar plates before isolating bacteria.  Alternatively, these items
can be purchased from Carolina Biological Supply or Wards Biological Supply.

Materials
The following recipe makes about 20 plates:
Equipment: 1 liter  Erlenmeyer flask
  balance, weighing boats and spoons
  aluminum foil
  hot plate or autoclave
  protective gloves
  20 sterile petri plates

Supplies: 11.75 g of Plate Count Agar
  tap water

Procedure
1) Weigh out 11.75 g of Plate Count Agar (PCA) and place in Erlenmeyer flask.

2) Add 500 ml of distilled water to the flask and mix thoroughly.  Cover with aluminum foil.

3) Boil contents on hot plate for 30 minutes, or autoclave at 121oC for 20 minutes.   Let cool for 5 minutes or until easy to handle.

4) With protective gloves, pour PCA mixture into 20 petri plates and let cool until solid (about 10 minutes)
NOTE:  Use caution when pouring plates.  To prevent contamination, only open the petri plate as long as absolutely necessary.

5) Label plate "PCA".  Allow the plates to dry out by placing them at room temperature for 48 hours.

Part I: Making the Extracts

Materials
Each group should be provided with the following:
Equipment: balance, weighing boats and spoons

Supplies: 1 ml alcohol in eppendorf tubes
  plant material of students' choosing

Procedure for Day 1
1) Weigh out 0.5 grams of material (or use volume measurement).  For comparative purposes, the same amount of
material should be used for each extraction.

2) Place weighed material into 1 ml of alcohol and allow mixture to sit overnight at room temperature. (This permits the
active compounds to move into the acetone)

_______________________________________________________________________________

Part II: A Drop of Healing
The instructor needs to have 1/4" sterile disks, and tetracycline antibiotic disks for this part.  These can be purchased from Fisher Scientific Supply.

Materials
Each group should be provided with the following:
Supplies: forceps
  alcohol in a small beaker
  1/4" sterile disks
  tetracycline antibiotic disk
  plastic or glass pipettes
  tooth picks
  sterile cotton swabs
  Plate Count Agar petri plates  (1 for each bacteria type tested)
  test bacteria (we used Staphylococcus epidermidis,   Eschericha coli and bacteria isolated from Compost)
 Bacterial strains can be obtained from Wards or Carolina Biological Supply.

Procedure for Day 2
Experiment
1) Obtain tubes from Part I.  Using the plastic pipettes, put one drop of the alcohol extract on a 1/4" sterile disk.  Allow disk
to dry in a clean, empty petri dish with the lid slightly open for at least 10 minutes.  If necessary, add another drop and allow it
to dry.

2) With a sterile cotton swab, spread each bacteria type onto a different Plate Count Agar plate (i.e. E.coli on one plate,
Strep. on another).
 NOTE:  Be sure to dispose of all contaminated objects to its proper container.
 To prevent contamination, only open the petri plate as long as absolutely necessary.

3) Soak forceps in alcohol for a few minutes and let air dry.

4) Using clean forceps, place your sample disks onto the Plate Count Agar from step 2
NOTE: Soak the forceps between each use.

Control
5) Prepare control disks by placing 1 drop of alcohol on a sterile disk and allowing them to dry.

6) Place a solvent control disk and a tetracycline-antibiotic disk onto the surface of each PCA plate from step 2, above.

7) Incubate the plates overnight at room temperature to allow the lawn of bacteria to grow.
____________________________________________________________________

Part III: Checking for Growth Inhibitors

Materials
Each group should be provided with the following:
 plates from Day 2
 ruler (with mm scale)

Procedure for Day 3
1) Measure the zone of clearing (if any) around the antibiotic disks.  A zone of clearing indicates  anti-microbial activity.  (Why?)
 

Do Soaps Really Do It?

Authors:
Kristen Wolfe, Kimberly Baldwin, Malik Williams, Sue Merkel

Abstract:
This activity allows students to explore how effective commercial soaps and detergents are, in relation to manufacturer claims that their soaps are "anti-bacterial". Students will screen different commercial soaps for the ability to inhibit bacterial growth.   Students develop their own experiments and controls to determine the effectiveness of a variety of soap products.  Most experiments will involve sampling a surface for bacteria, then using different soaps on their skin or an inanimate surface, and then resampling to test for the presence of bacteria.  This lab activity was designed for and tested in middle or high school biology classrooms.

Introduction:
There has been increased interest in bacteria that lurk everywhere in our homes and kitchens, ready to attack and cause disease.  Many soap manufacturers have jumped on the band wagon by providing products that claim "new and improved anti-bacterial activity."  But do these products really work?  Here is a chance for students to test and compare different products for themselves, by sampling a surface, washing it with soap, then sampling again.  Most students will find ambiguous results after the first day because: they did not have the proper controls, they did not sample equal areas, or the swab was wet one time, but not the other.  This lab gives students a chance to redesign their labs to address the ambiguities, and illustrates why controls are so critical.

Special Materials:
Equipment:
Supplies: Petri plates with Plate Count Agar
 sterile cotton swabs
 a collection of commercial soap products
 
Time Required
Prep Day Prepare the petri plates with Plate Count Agar.
 
Day 1 talk about bacteria and introduce students to  Plate Count Agar petri plates.
 Have students formulate a hypothesis about one r more soap product, (does it really kill bacteria?) and design
an experiment to test that hypothesis using cotton swabs and PCA plates.  Carry the experiment out.

Day 2 Observe the PCA plates.  Talk about student experiments and controls.  Have student redesign their experiments,
based on the discussion. Carry out new experiment.

Day 3  Observe the PCA plates.
 

Prep:  Making up the Plate Count Agar Medium
Instructor or students need to make Plate Count Agar plates before isolating bacteria.  Alternatively, these items can be purchased from Ward's Biology: 1-(800) 962-2660 or Carolina Biological Supply: 1-(800) 334-5551)
 

Materials
The following recipe makes about 20 plates:
Equipment: 1 liter  Erlenmeyer flask
  balance, weighing boats and spoons
  aluminum foil
  hot plate or autoclave
  protective gloves
  20 sterile petri plates

Supplies: 11.75 g of Plate Count Agar
  tap water

Procedure
1) Weigh out 11.75 g of Plate Count Agar (PCA) and place in Erlenmeyer flask.

2) Add 500 ml of distilled water to the flask and mix thoroughly.  Cover with aluminum foil.

3) Boil contents on hot plate for 30 minutes, or autoclave at 121oC for 20 minutes.   Let cool for 5 minutes or until easy to handle.

4) With protective gloves, pour PCA mixture into 20 petri plates and let cool until solid (about 10 minutes)
NOTE:  Use caution when pouring plates.  To prevent contamination, only open the petri plate as long as absolutely necessary.
 
5) Label plate "PCA".  Allow the plates to dry out by placing them at room temperature for 48 hours.
 

Part I: Asking a Question

Materials
Each group should be provided with the following:
 soap products
 2-4 PCA plates
 sterile cotton swabs

Procedure for Day 1
1) Discuss what bacteria are and how we try to control growth with soaps.  Have students develop a hypothesis about
one of the soaps, (EXAMPLE: Clean-O soap kills 100% of all bacteria in 30 seconds!); and develop an experiment to
test the hypothesis (EXAMPLE: sample skin, apply Clean-O for 30 seconds; sample skin again).  Skin and surfaces
can be sampled by rubbing them with a sterile cotton swab, then rubbing the swab on the surface of the PCA plate.

2) Carry out the experiments, and incubate PCA plates overnight (or more).
_______________________________________________________________________________

Part II: Checking Results and Redesigning the Experiment

Materials
Each group should be provided with the following:
 soap products
 2-4 PCA plates
 sterile cotton swabs

Procedure for Day 2
1) Observe the plates.  If your results are ambiguous, redesign the experiment to better answer your original question.
Think about what controls you had, and which ones you may need.

2) Carry out the experiments, and incubate PCA plates overnight (or more).

3) Observe the plates and summarize your results.

Genetic Exchange in Bacteria

Authors
Susan M. Merkel and Adam M. Chazan
Section of Microbiology, Cornell University, Ithaca, NY

Abstract
This activity demonstrates the exchange of genetic information by bacterial cells, using competent E. coli cells and a plasmid carrying a gene for Green Fluorescent Protein (GFP).  We describe a simplified method for making competent E. coli and for carrying out the transformation.
Although this process is very common in nature and in research labs, it is not always easy to observe.  This obstacle is overcome by using the GFP as a marker to visualize gene movement.  Not only is GFP very easy to detect, it is quite striking
to observe.  This lab is meant for very advanced high school classes, or introductory college level classes.
 

Introduction
The exchange of genetic information (DNA) in bacterial cells has helped to drive evolution since life began.  In more
recent times, scientists have developed tools that allow us to purposefully move genes from cell to cell, paving the way for
recombinant DNA technology.   There has been a renewed interest in and concern about these processes, as the exchange
of DNA among bacteria contributes to the rise in antibiotic resistance observed lately.

Special Materials
Equipment: tabletop centrifuge
 20 - 200 ul pipettors

Supplies:  a special strain of E. coli, carrying the GFP plasmid
 a non-GFP strain of E. coli
 ampicillan
 100% ethanol
 RNAase
 plastic eppendorf tubes
 

Time Required
Prep Day For the plasmid preparation: Make TE buffer and Solution and III (see Appendix for recipes).
Grow E. coli (GFP) in 4 ml LB+AMP broth at 37oC overnight.

 For making competent cells:

Day 1 For the plasmid preparation: Prepare a fresh solution of NaOH/SDS solution.   Centrifuge the cells,
and extract the plasmid DNA.  Rinse with EtOH.  Store in TE buffer.

 For making competent cells:

Day 2:  Carry out the genetic exchange through transformation.  Spread cells on LB Agar plates.  Incubate at 37oC overnight.

Day 3: Look for transformed colonies.
  

Preparation: Making solutions and growing cultures

Materials
Equipment: incubator
Supplies: 5 M potassium acetate
 Glacial acetic acid
 

For the plasmid preparation:
 Make TE buffer, GTE Solution and Acidic Solution.  Grow E. coli (GFP) in 4 ml LB+AMP broth at 37oC overnight.
(See appendix for medium recipe)

TE Buffer
1) Prepare 1 l of a 25 mM Tris-Cl (pH 8.0), and 10 mM EDTA (pH 8.0) solution.

GTE (Glucose/Tris/EDTA)  Solution
1) Add 50 mM glucose to 100 ml TE buffer.   Autoclave for 15 minutes; store at 4oC.

Acidic Solution
1) CAREFULLY mix together 60 ml of 5 M potassium acetate, 11.5 ml of glacial acetic acid, and 28.5 ml of H2O
 

Overnight Culture of E.coli (GFP)
NOTE: cultures of E.coli (GFP) should be grown in the presence of 50 ug/ml ampicillan, to help maintain the plasmid
in the cells.

1) Transfer a single colony of E.coli (GFP) into 4 ml (LB + 50 ug/ml ampicillan) broth. Shake (if possible) overnight at 37oC.

For making competent cells:
TSS buffer

Overnight Culture of E.coli ( no GFP)
1) Transfer a single colony of E. coli (no GFP) into 4 ml LB broth. Shake overnight at 37oC.
 

Part I: Plasmid Preparation

Materials
Equipment: vortex (if possible)
 table top centrifuge

Supplies: NaOH solution
 Acidic solution
 TE buffer
 GTE solution
 glacial acetic acid
 100% ethanol
 bucket of ice

Procedure for Day 1

Make NaOH/SDS Solution
1) Dilute 10N NaOH to a concentration of 0.2 N Na OH in 1% SDS (for example, add 0.5 ml 10N NaOH to 25 ml 1% SDS).  Prepare fresh immediately before use by mixing ingredients in a test tube, and inverting 5 times (do not vortex).
Store tubes on ice.
 

Plasmid Preparation
1) Place GTE and Acidic solutions on ice.

2) Pour 1.5 ml of the overnight cultures into eppendorf tubes. Centrifuge at 12,000g for 1 minute (top speed on table top centrifuge). Remove broth and repeat.

3) Resuspend pellet in 100 ul ice cold GTE Solution.  Vortex or shake by flicking tube with index finger.

4) Add 200 ul NaOH/SDS solution (freshly prepared).  Invert tubes several times to mix.  (This lyses the cells.)

5) Add 150 ul ice cold Acidic solution.  Close tube and invert for 10 seconds.  Place on ice for 5 minutes. (This helps to separate the double stranded DNA.  The large chromosome can't get back together; the smaller plasmids can.)

6) Centrifuge at 12,000g for 15 minutes.  Transfer supernatant to fresh tube.
(The smaller plasmids should be dissolved in the supernatant.)

7) Precipitate DNA with 2 1/2 times volume cold ETOH. Vortex, and place on ice for 20 minutes.

8) Remove supernatant and allow pellet to dry by inverting over a paper towel.  (The plasmid DNA should appear
as a small white dot on the side of the tube.)

7) Rinse with 1 ml cold 70% EtOH, being careful not to disturb the pellet.  Air dry again.

8) Dissolve the pellet in 50 ul TE buffer.  Store at -20oC.
 
 

Part II: Making competent E.coli cells

Materials
Equipment: 1 liter Erlenmeyer flask
 floor centrifuge/ centrifuge tubes

Supplies: TSS buffer
 bucket of ice

Procedure for Day 1
1) Beginning early in the day, place 1 ml of the overnight culture into 100 ml LB broth (use a 1 liter Erlenmeyer flask).
Shake at 37oC until solution is cloudy (absorbance  = about 0.3; this may take 3-4 hours)

2) Spin 20 ml culture at 10,000g for 10 minutes.

3) Pour off the supernatant.

4) Add 2 ml cold TSS buffer.  Mix gently to resuspend the cells.  Place on ice.
Cells are now ready to take up DNA or can be stored at -70oC.
 
 

Part III: the Transformation (genetic exchange)

Materials
Equipment: 37oC incubator
 timer

Supplies: eppendorf tubes
 bucket of ice
 10 mls LB+ 20 mM glucose broth

Procedure for Day 2
Experiment
1) Place 100 ul competent cells in sterile eppendorf tubes.  Place on ice.

2) Add 20 ul plasmid DNA solution to tube

3) Incubate on ice for 30 minutes.

4) Incubate at 37oC for 20 seconds.

5) Incubate on ice for 2 minutes.

6) Add 1.9 ml prewarmed LB+ 20 mM glucose broth to each tube. Incubate 60 minutes at 30oC.  Shake.

7) Plate 0.1 ml cells on each of 5 - 10 LB (+ampicillan) plates.  Incubate overnight at 30oC.

Controls
8) Plate 0.1 ml untransformed, competent E.coli (no GFP) cells on one LB (+ampicillan) plate.  Incubate overnight at 30oC.

9) Plate 10 ul of the plasmid preparation on one LB (+ampicillan) plates.  Incubate overnight at 30oC.

Procedure for Day 3
1) View all the plates under long UV radiation to see if any cells "picked up" the plasmids DNA with GFP gene.  These colonies should glow bright green under the UV lamp.

Appendix

 Culture Media

Plate Count Agar (per liter)    Starch-Casein Agar (per liter)
5g    Bacto Tryptone      10g    Soluble Starch
2.5g Bacto Yeast Extract     0.3g   Vitamin free casein
1g    Bacto Dextrose (glucose)    2.0g   Sodium Nitrate
15g  Bacto Agar      2.0g   Sodium chloride
1000 ml of water      2.0g   Dipotassium phosphate
0.05g Magnesium sulfate 7H20
0.02g Calcium carbonate
0.01g Ferrous sulfate/ 7H20
15.0g Bacto agar
1,000 ml Tap water
LB Broth (per liter)
10g Bacto Tryptone
5g   Bacto Yeast Extract
10g Sodium Chloride
1000 ml water
 

Supply Houses

Ward's Biology: 1-(800) 962-2660

Carolina Biological Supply: 1-(800) 334-5551

Fisher Scientific Company 1-(800) 766-7000
 
 

MicroBiology Ambassadors-Fall 1997  page