BIOLOGY 2120             CELLULAR AND MOLECULAR LABORATORY                                      FALL 2000

                                                                               POLICIES

Organization:  Each lab section meets once a week as shown on the syllabus.  Readings will be from the laboratory exercise book and any readings specified therein.  Read the material for each exercise before you arrive in the laboratory.  If there are supplementary handouts, they will be distributed at the Biol. 2320 lecture on the Monday of the lab exercise.  The recombinant DNA labs (weeks of 10/2, 10/9, 10/16 & 10/123) will use Karp pages 795-796, 800-820, & 811-812 a supplementary references.  We will also be using lab time to discuss topics being presented in lecture.

Attendance:  Attendance is required at all scheduled labs.  Do not assume that labs will finish on time.  There will often be a discussion of results at the end of lab.  Do not leave the lab until this discussion has taken place, or until the instructor indicates that it will occur at the beginning of the following lab.  Absences may be made up in the same week by attending the other lab section, but only by prior arrangement with the instructor whose section you intend to attend.  There are also several types of excusable absences (adequate documentation of illness, family business, official university functions such as varsity athletics) for which there will be some type of make-up process.  This may include getting data from another student and completing the homework from that data, or it may simply be letting you take a test a week late.

Appropriate lab attire and accessories: Choose clothes to wear on the assumption that they may get stained or damaged, and thus may protect you from the same fate.  Do not wear dangling jewelry or loose clothing.  These could get caught in electrical or mechanical instruments; this could kill you.  For the same reason, tie back long hair.  Closed-toe shoes are required.  Food or drink is not an appropriate accessory unless specifically approved for that week.

Grading:  You will be graded on the following types of coursework:

               Tests (approximately every other week, 5 in all)                                                                              50%
                         Worksheets, homework and participation                                                               10%
                         Writing assignments                                                                                                   40%  

Individual instructors may adjust weighting of components.  Tests are scheduled as indicated by the asterisk (*) next to the lab dates on the following pages.  Tests will cover the preceding labs not previously tested on, and may include a question concerning the current week's lab exercise.  Worksheets and homework are to be turned in as required.  For the writing assignments, specific labs will be chosen as the subject of a written description of one or more parts of the lab. These compositions will be written as if to appear in a scientific journal (Introduction, Materials and Methods, Results, and Discussion). The final assignment will require a complete paper in this format. Detailed instructions on how to do this will be provided and discussed during lab sessions.

                                                                          SCHEDULE

                   CELLULAR AND MOLECULAR BIOLOGY-LABORATORY (BIO 2120)

Week               Topic
of:  

               8/28                  Organization and basic techniques

               Review of dilutions and spectrophotometry plus basic practice with pipetting and sterile technique.  See Procedural note below.

               9/4                    DNA Isolation as an example of Cell Fractionation

               Total DNA will be isolated from E. coli, and evaluated for purity and yield by spectroscopic methods.

               9/11*                Enzyme kinetics

                                         The Michaelis constant for purified ß-galactosidase will be determined.  

               9/18                  Separation of proteins by size and charge (And the Biorad protein assay)

               A range of proteins will be separated by charge and molecular weight will be estimated using a pair of electrophoretic techniques: native and SDS gels.  The Biorad protein assay will be performed as an example of the use of standard curves.

9/25*               Library exercise/Discussion of writing assignments

                                         Science librarian will instruct students in use of reference facilities.  

10/2                  Recombinant DNA I - Molecular Cloning

               DNA from cheek cells will be obtained, and a repetitive sequence amplified by polymerase chain reaction.  Agarose gel electrophoresis will be performed on standard fragments of DNA to introduce procedures for molecular size determination and mapping via restriction endonuclease digestion.

                10/9                  Recombinant DNA II - Molecular Cloning

                The presence of a fragment of the LINE-1 repetitive sequence, from the PCR amplification reaction will be confirmed by agarose elctrophoresis.  The fragment will then be purified and ligated into a cloning vector.  The ligation products will be used to transform E. coli, and transformants will be scored for the presence of recombinant plasmid.

               10/16                Recombinant DNA III - Plasmid Purification

               The plasmid carrying LINE-1 will be purified by the alkaline lysis technique and restriction endonuclease digests will be performed.

10/23*             Recombinant DNA IV - Sequence Analysis

               The LINE-1 sequence will be analyzed by restriction endonuclease digestion and fragment length determination by electrophoresis.

10/30                Reading the scientific literature

                         One or more papers will be analyzed during lab, and a follow-up assignment given.

               11/6                  Permeability and chemical properties of red blood cell membranes

               The cell lysis method will be used to determine permeability and osmotic properties of red blood cell membranes.  The effects of reagents altering membrane structure will also be examined.

               11/13*              Photosynthesis I: electron transport as energy transduction

               Chloroplast membranes are prepared for quantitative measurement of electron transport.  The correlation between energy input and rate of reduction of DPIP will be determined

               11/27                Photosynthesis II: chemosmosis

Chloroplast membranes including sealed thylakoids are prepared for measurement of light-induced alkalinization of the medium around the membranes as an indicator of chemosmotic energy transduction.

               12/11*              Last Quiz (Common exam period I, 6.30 PM, 12/11)

This is not a final exam, simply a quiz over the bioenergetics/photosynthesis labs.  Tests for all sections will be on the same day, but different instructors will give different tests.

Procedural Note:

                          There will be a hands-on practical quiz on dilutions and pipetting after the first lab.  Instructors will schedule 15 min quiz times from 9/6 - 9/20.  Each student takes the quiz separately.

Editorial Note:

                         There are some places where the text indicates that you are to graph data "on the next page".  In these instances, graph paper will be provided in the laboratory.  Make sure the graph you include presents the data properly.  Check with your instructor if you are not sure.