Bios 250A Research Project Fall 1999

(fornari)

Introduction

After learning and practicing the four basic methods of Microbiology, each lab group will embark upon a group-designed project (i.e., the members of the group completely design, execute, evaluate and present the history, data and results of the project). The lab research project is scheduled for the last half of the semester, and will culminate in a series of presentations by the members of each group. I will continue to enter each week=s lab schedule on the Web-site syllabus, especially since your project and the biochemical and genetic testing labs will overlap.

Objective

The project=s objective is to thoroughly evaluate and identify a microbial by both biochemical and genetic methods. At least as many different microbial species as there are members in the group must be identified and analyzed (e.g., a 4 member group must identify and analyze 4 different microbial species). The main objective includes not only identifying the genus and species, but also becoming thoroughly familiar with all the ecological, environmental, physiological, biochemical and genetic properties of your microbial species

You will be assigned an unknown microbe, which will be distributed on an agar slant. This slant is your main stock, and you must make a “working” stock from this slant. The working stock is the one you will use to perform all your lab tests and analyses. When you receive your slant, immediately begin your research project by noting the characteristics of your strain growing on the surface of the agar. You are starting your project in the “inductive” phase of the scientific method, by noting all the particulars of your unknown strain. Once you have collected sufficient information from experimental tests (see below), you will analyze this data for a pattern. This pattern will allow you to generate an initial hypothesis about the identity of your unknown. More tests will then be run to confirm or refute your initial guess or hypothesis. Remember that a “good” hypothesis has built into it a number of testable predictions.

Means to achieve your Objective

After making working stocks (see lab exercises 31 & 32) of each bacterial type from streak plates (see also the flow-chart), your first set of experiments to identify and characterize your unknown strains will of course be Gram-stains (see also exp. # 31. The next set of experiments will be biochemical in nature, and will be performed along with known controls provided to you in the lab. Following a battery of selected biochemical tests, you will have an opportunity to use the Bio-Log system, in which biochemical tests are matched against a computer database of information about bacterial species.

The use of Bergey=s Manual of Determinative Bacteriology (and other sources which I will provide on a separate handout) will aid you in making the final identifications of your site-isolates. In some cases you may not be able to identify the species and will have to settle for a genus identification only. In these (rare) cases, you should make every effort to provide at least some good estimations of the possible species that seem to best fit your collected data, i.e. you will have multiple hypotheses.

This semester we will also attempt to perform molecular genetic analyses of the DNA from each unknown. Rapid methods of isolating the DNA will be used, and the isolated DNA will be analyzed by the Polymerase Chain reaction (PCR), followed by gel electrophoresis. You should view these molecular tests as confirmations (or denials) of your hypotheses.

A large variety of lab manuals containing an even lager variety of experiments is available to you in the lab. These manuals with their exercises and tests are good sources of both ideas and new experimental tests you might want (need) to perform in order to identify your site-isolates. In most cases of applying any new tests, special supplies and chemicals will be needed; after carefully reviewing what you might need in order to perform a particular test, you will inform your lab T.A. of these items so she can arrange for them to be procured or made by the student media makers under Jason Lemberg=s supervision. Please allow sufficient time in between a request and its delivery to you; careful planning on your part is essential for a timely accomplishment of all your experiments.

Report and Presentation

Upon completion of your research project, you will write a group report (with all members signing the final report; sample reports will be provided) and prepare a presentation for the class. The formats for the report and the presentation will be discussed in lab and on another hand-out. In the interim, it is absolutely essential that all members in each lab-group maintain accurate and detailed records of all experimental protocols and collected data. Such records and data will make it much easier to write the final report, and assemble all data and information into over-heads (or Power-Point “slides”)for your class presentation.

Summary of Research Project Plan:

I. Use of Classical Microbiology techniques and methods (4 basic techniques)

II. Selected Biochemical tests

III. Bio-log computer assisted analyses

IV. PCR molecular genetic analyses

V. Report and Presentation

Your final, completed project will be some unique mix of the above approaches, coupled with a thorough understanding of the properties of your identified microbe. This project involves as much “library research” as is does lab work.