ESTCP Project #1011, Rhizosphere
Final Report
3.26 g BH medium
1.0 L deionized H2O
(one drop cyclohexanol (Aldrich#10,589-9) added to each of 40 test tube
containing 4.5 mL BH medium/tube)
Materials (for one soil sample)
170
test tubes (6-10mL)
5
test tube racks
170
test tube caps
Aluminum foil
1
multi-channel pipetter
40
sterile tips for multi-channel pipetter
8
sterile multi-channel pipetter basins
1
aluminum weigh dish
25
mL BH negative control medium
25 mL
Dextrose positive control medium
200 mL
Motor Oil medium
200 mL
Vegetable Oil medium
200 mL
Sodium Benzoate medium
200 mL
Cyclohexanol medium
95 mL MPP dilution bottle (10-1 dilution)
1
90 mL MPP dilution bottle (10-2 dilution)
1
45 mL MPP dilution bottles (10-3 through 10-9 dilutions)
7
(total of 9 MPP dilution bottles per sample)
9
stoppers for dilution bottles
1
wide tip 10-mL sterile pipette
7
5-mL sterile pipettes
1
pipette bulb
MPN Media Preparation per Sample
1. Prepare the MPP buffer using direction from Appendix B. You will need 9 dilution
bottles and 9 stoppers per soil sample.
2. BH negative control: Prepare medium using recipe on media page, add 4.7 mL of
medium to each of 5 test tubes. To allow for volume loss during autoclaving, initial tube
volumes should be measured to 4.7 mL to achieve final volumes of 4.5 mL of medium.
The 10-2 dilution will be the only dilution inoculated for BH negative control medium.
Cover test tubes with foil and autoclave @ 121C for 15 min.
50
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