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Screening for Active Ingredients in Plant Extracts that Inhibit the Growth of Agrobacterium tumefaciens

1. Agrobacterium tumefaciens inoculum may be ordered from several commercial laboratories. We purchase ours from Presque Isle Cultures, Erie, PA. Website: http://www.picultures.com
2. The initial lab exercise can be completed in about one hour. A follow-up lab session (about 1 hour) is used to record and discuss observations.
3. You may wish to slice the carrot disks as the students are setting up for the experiment. It saves time and ensures uniform-sized disks. If the disks are too large, the lid of the petri dish will smash the top of the carrot disk and interfere with gall formation.
4. We found it necessary to teach students how to correctly use Parafilm to seal the petri dishes.
5. The petri dishes were placed on a lab bench under ambient lighting conditions through the duration of the experiment.
6. Contamination from bacteria or fungal spores due to nonsterile techniques will show up quickly, usually within several days. In petri dishes that were not contaminated, it took one to two weeks to see the beginning of gall formation on disks that formed galls.
7. Only two of our students screened plants materials that appeared to inhibit growth of A. tumefaciens, but we discussed in a subsequent lab the fact that those treatments would need to be replicated under more carefully controlled conditions to determine if they really were effective or if an experimental error gave false results.
8. This experiment could be easily adapted to screen for inhibitors of other plant pathogens. Students could also screen with materials such as spices or 'natural' household cleansers.

Instructors may read suggested answers to discussion questions in the password-protected area of the Instructor Communication and Scholarship section of this site, which can be accessed at http://www.apsnet.org/education/InstructorCommunication/ExamQA/Top.html