Procedure 

Materials

• Meter stick
• 80 metal flags (to mark corners of plots and to hold down screens)
• Small-holed black mesh
• Unpainted canvas
• Hard black plastic (i.e. folders)  
• Saran wrap
• Easygel™ petri dishes
• Plastic test tubes with caps
• Test tube racks
• Distilled water
• Transformation tubes
• Individual bottles of Easygel™
• Pipette (10 mL) and pumps
• Micropipette pumps (100 µl)
• Micropipette tips
• Tip sterilizing solution (10% bleach solution)
• Permanent black marker
• Glass stirring rod
• Plastic bags
• 1 cm³ measure

Procedure:

1)    Cut 4 15 cm x 15 cm squares of a mesh with fairly small holes

2)    Repeat step 1 for hard black plastic, plastic clingwrap, and unpainted canvas

3)    Cut small slits in each corner of the squares

4)    Label 80 flags with Site, Quadrat, and Screen type (i.e. Site 4, Quadrat 1, Mesh).  There should be 4   flags for each plot of land  

5)   Select a location with maximum sunlight in each Quadrat of site 4 and one in site 1, and clear a square half-meter of soil in each location (removing or bending back all plants and removing all dead leaves, woodchips, etc. so that the soil is completely clear of all debris)

6)   Go to the cleared plot in Quadrat 1

7)    Place a plastic clingwrap square over a portion of the cleared area and place the “Site 4, Quadrat 1, Plastic clingwrap” flags at each corner of the plastic clingwrap square through the cut slits, securing it firmly to the ground

8)    Repeat step 7 for mesh, canvas, and black plastic

9)    Repeat steps 7-8 in Quadrats 2, 3, and 4

10)    At the cleared plot in site one, repeat step 7 four times in the northeast, southeast, northwest, and southwest corners. 

11)    Label 20 plastic bags like the flags according to site, cover, and Quadrat (i.e. Site 4, Quadrat 1, Canvas), and label 15 more bags for Quadrats 2, 3, and 4 and Site 1, northeast, southeast, and southwest

12)   Wait 24 hours (until day two of experiment)

13)    At plot in Quadrat 1, take a 10 cm deep soil core sample from the exact center of the plastic clingwrap cover and place in the appropriate plastic bag  

14)    Repeat step 13 for each cover (mesh, canvas, and hard black plastic)

15)    In Site 1, repeat step 13 for the northwest sample under the plastic clingwrap

16)    Label 105 Easygel plates with Site, Quadrat, Cover, and Dilution Level (i.e. Site 4, Quadrat 1, canvas, 10^-2)

17)    Perform SERIAL DILUTIONS to 10^-4 for each soil core sample taken and PLATE the 10^-2 to the 10^-4 dilutions on appropriate Easygel™ plates

18)    Let remaining samples sit until Day 5 of the experiment and repeat steps 13-17 for Quadrats 2 and 4 in Site 4 and the northeast and southwest samples in Site 1 

19)   Count the number of molds growing on Easygel™  petri plates that were plated on Day 2 of the experiment

20)    Let remaining samples sit until Day 6 of the experiment and repeat steps 13-17 for Quadrat 3 in Site 4 and SE in Site 1

21)    On Day 7 of the experiment count the number of molds plated on Day 5

22)    On Day 8 of the experiment count the number of molds plated on Day 6

23)    Take all data and use the equation: # of colonies * 10⁴ * 10 ^{|dilution level of sample|}    to calculate the # of mold per cm³ of soil and then average the data from each cover taken on a given day

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