BISC209/S13: Lab10

Lab 9 LAB PRACTICAL

Today you will have a lab practical exam designed to assess your mastery of basic tools, techniques, and theoretical information on which the field of microbiology is based.

Graded Assignment: Make separate figures and text for the antibiotic assay (if any of your site teammates indicated the presence of an antibiotic producer) and interactions assays. Use the Google doc table of the tests performed on the isolates from your soil community (include your teammates' isolates from the same sampling site for this assignment) to make a Table and write the results narrative for this table. You should include in the table: Gram stain, enrichment/selection medium, description of the colony morphology, description of the individual bacteria shape (cocci, bacilli, spirilla) and any useful descriptors ie., large, small, bullet shaped, in chains, etc., evidence of endospores (either endospore stain positive or visualization of empty areas in vegetative cells on Gram stain), & motility. Include metabolic tests such as nitrate reduction and mannitol fermentation. Leave out the interaction assay and the antibiotic producers test (if your site had a positive) in this table and make separate results figures from photographs of those plates and discuss those findings in separate sections in results. Work with ALL of your sampling sites' interaction assays on this results section and a photo of any positive antibiotic producers from your sampling site for that section of this draft results. If your site had no positive antibiotic producers, you can include this test and its negative results in your table with out a photo or separate section.

The results narrative for the table of tests performed on the isolates might start out, "In order to show structural and metabolic diversity as well as the potential for community behavior (co-operation and/or competition) among members of a soil community, nitrogen cycling bacteria & spore-forming bacteria were enriched and selected using media and protocols described in Materials and Methods. Once in pure culture a few soil community members of these groups and other interesting bacteria isolated on non-selective media were visualized using staining techniques for cell wall structure in a Gram stain, production of endospores with Shaeffer-Fulton endospore stain, the presence of capsules with a negative nigrosin stain, and flagella with a flagella stain. Additionally motility testing was performed on each isolate using soft agar deeps. Evidence for a role in the nitrogen cycle was assessed by________. (continue to describe any other testing you include in your table)......Table 1 shows the results of this testing."

After this results section introduction, you can then analyze the findings in a narrative with the goal of using the evidence in the table to answer our experimental question, "is there evidence for diversity and functional co-operation and competition among the bacteria isolated from your soil community?"

This lab be devoted to learning how to identify bacteria by 16s rRNA gene sequences and to instructor designed workshop activities to help prepare you to write your final paper.

Assignment

(If your group's gene sequencing didn't work well, omit the exercise described in this paragraph.)If your soil sample's 16s rRNA gene sequencing yielded enough data, prepare draft table/figure(s) on the information gleaned from your RDP data base analysis of the DNA sequence information on the cultured isolates from your soil sample. This analysis should identify them as much as possible (from the 16s rRNA gene sequence comparisons done using the RDP data base or from an NCBI BLAST. Do your isolated bacteria belong to the genus or family you expected from the morphologic and metabolic information you have saw in your test results? You can use Wellesley's electronic version of The Procaryotes found at [1] and/or Bergey's Manual found at [2] to look up more information on the isolates identified by 16s rDNA sequencing. It would be interesting to compare the characteristics described in these reference books to the characteristics you noted in your analyses. Do you have evidence for phylogenetic diversity in your soil sample? (This is a different form of richness in the community.) Create a phylogenetic tree of your soil sample's community members (using a variety of non-members as reference points to try to determine evolutionary relatedness among your soil sample's cultured isolates. Keep in mind that if you find broad phylogenetic diversity it is significant and can be used as evidence for another kind of richness in your microbial soil community. However, if all your cultured isolates belong to closely related clads, it does NOT mean that your soil community is not phylogenetically diverse. Why?

Your research report on your semester project is due in your lab instructor's office by 5pm on May 7 or May 8 . Please make sure there is an electronic copy of your paper uploaded to your folder in your Lab's Drop Box in Sakai by that time. The electronic copy is required in addition to a required hard copy that must be submitted to your instructor per her instructions.

More information about the final paper can be found at: Assignment: Assignment: Final Paper. A grading rubric is posted to the Rubrics folder in the Resources section of your lab Sakai site.

Discard and clean up any remaining tests and cultures on your isolates. Note that there are 5 bonus points awarded for perfect clean-up. Your instructor will explain the "rules" for obtaining these bonus points.

Links to Labs

Lab 1
Lab 2
Lab 3
Lab 4
Lab 5
Lab 6
Lab 7
Lab 8
Lab 9
Lab 10