University of Iowa
Department of Biochemistry
4-403 BSB
Iowa City, IA 52242-1109 USA phone: 877-846-8569
or 319-335-7932
fax: (319) 335-9570
biochem@uiowa.edu

Department of Biochemistry

99:140 Experimental Biochemistry

Instructors: David Price (course director), 3130 MERF, 335-7910, david-price@uiowa.edu; Elisabeth Swain, 4-552 BSB, 335-8569, elizabeth-swain@uiowa.edu; Todd Washington, 4-734 BSB, 335-7518, todd-washington@uiowa.edu; Christine Kondratick, 4-700 BSB, 335-7501, christine-kondratick@uiowa.edu

Time & Location:
Lecture: Friday, 2:30-4:00 PM, 1107 BSB
Lab section A01: Monday, Wednesday, 1:30-5:00 PM, 221 MEB
Lab section A02: Tuesday, Thursday, 1:30-5:00 PM, 221 MEB

Hrs: 4 s.h.

Description: Use of modern instruments and techniques to fractionate, identify and characterize constitutents of biochemical systems. Prerequisite: 099:120 with grade of C or above; 99:130 (may be taken as corequisite). Corequisite: 099:130, if not taken as prerequisite.

Course Requirements: completed lab protocols prior to each lab session; laboratory report for each course unit (four); four examinations. Laboratory attendance is mandatory.

Syllabus, spring semester, 2008

Text: No textbook. Instructor provides course material for each lab exercise.

LECTURE SCHEDULE :
Check in / Lab safety video
UNIT ONE
Lecture: Protein Purification
Lab: Preparation of cherry cotyledon extract
Lab: Con A Chromatography of Amygdalin Hydrolase
Lecture: Electrophoresis of Proteins
Lab: Ion exchange chromatography
Lab: SDS polyacrylamide gel electrophoresis
Lecture: Analysis of the Kitetic Properties of Enzymes
Lab: Specific Activity Measurements / Western blot
Lab: Km, Vmax, / Western blot
Lecture: RNA Polymerase II Elongation Control and Recombinant Protein Expression
Lab: Inhibition Study
FIRST EXAM
UNIT TWO
Lab: Analysis of the RNA binding properties of human HEXIM1
Lab: Restriction analysis, transformation and plating of E. coli
Lecture: Binding of Proteins to Nucleic Acids -- HEXIM1
Lab: Bacterial expression and purification of HEXIM1: Expression
Lab: Bacterial expression and purification of HEXIM1: Analysis
Lecture: none
7SK RNA: Template preparation and in vitro transcription
Mobility shift assay: Analysis of HEXIM1 • 7SK complex
Lecture: none
Mobility shift assay: Competition binding
Review
SECOND EXAM
UNIT THREE
Lab: Virtual cloning (electronic classroom)
Lab: Prepare genomic DNA and PCR amplification
Lecture: Recombinant DNA, part 1
Lab: Restriction digest and analytical gel electrophoresis
Lab: Isolation of DNA fragments from a preparative gel and DNA ligation
Lecture: Recombinant DNA, part 2
Lab: Making competent bacterial cells and transformation
Lab: Plasmid preparation, analytical gel, DNA sequence analysis
Lecture: DNA Repair and Mutagenesis in Eukaryotes
Lab: Viewing protein structures; DNA sequencing results (electronic classroom)
THIRD EXAM
UNIT FOUR
Lab: Site-directed mutagenesis I
Lecture: Eukaryotic vectors
Lab: Site-directed mutagenesis II
Lab: DNA sequencing
Lecture: UV Sensitivity and Mutagenesis Assays
Lab: Prepare mRNA, RT-PCR
Lab: UV sensitivity and mutagenesis assays
Lecture: Protein Expression Systems
Lab: Analysis of UV experiments and RT-PCR products
Lab: Lab clean-up and review
FOURTH EXAM