Dr. John H. Williamson
Department of Biology
This course will emphasize research techniques
in biochemistry. The first part of the course will be highly structured,
learning techniques and how to "do" research. The primary goals
of the course will be:  "hands on" experience,  "how"
to ask questions,  the use of "research equipment" and 
"communication" of scientific information. The central theme connecting
these goals will be the purification and characterization of NADP+-isocitrate
dehydrogenase [IDH] from an organism of specific interest. Techniques to
be learned will include spectrophotometric enzyme assays, protein assays,
centrifugation, polyacrylamide gel electrophoresis, gel filtration chromatography,
affinity chromatography, data management, computer graphing and the writing
of a manuscript to be submitted for consideration of publication. The final
"technique" will be the presentation of a research poster at an
"open house" for the Biology Department.
I expect students in this course to be mature,
independent, highly motivated self-starters who are willing to work as a
team. You should recognize, at the beginning, that research projects do
not fit a "3-hour lab session on a one day per week schedule".
Thus, we will need to determine a schedule that we can all meet and you
will be expected to be there, having "done your homework" and
prepared to make meaningful contributions. In addition, all of us will be
required to come in at "off schedule" times to run or complete
You will receive weekly assignments that will be marked and returned immediately. Your contributions to weekly goals and plans will be evaluated and discussed with you. Suggested improvements, as appropriate, will be forwarded to you. Your timely management of experimental results, ie., immediately after data collection, will be essential to planning the next experiment or, indeed, whether the current experiment should be repeated or re-designed. A computer with a graphing program will be designated to the lab for your use. The breadth and quality of your contributions will be a major component of your grade for the semester.
Week number and Technique
1 Spectrophotometry / Enzyme Assays
2 Cellular Fractionation and Centrifugation
3 Affinity Chromatography: A Two Step Method
4 Affinity Chromatography: A Three Step Method
5 Protein Determination: Lowry Method
6 Enzyme Kinetics: Km - Substrate
7 Enzyme Kinetics: Km - Coenzyme
9 Temperature / pH Profiles
10 Divalent Metal Requirement
11 Polyacrylamide Gel Electrophoresis
12 - 14 Research Projects
15 Poster Presentations (Poster 1) (Poster 2)
YOUR FIRST ASSIGNMENT
Go to the library and, using Med-Line and / or other databases, search the literature since 1993 for publications directly concerning NADP+-dependent IDH.
If our library has an important paper, I will make a copy for all of us. If not, I will order it through interlibrary loan.
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College, Davidson, NC 28036
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