INSTRUCTOR:  Barry Honda, SSB 7140.

PREREQUISITES:  BISC 331, or permission of instructor. Enrollment may have to be limited.

DESCRIPTION:

The lectures will cover recombinant DNA methods, around a framework such as:

Prepare foreign DNA
Prepare vector
Join the two ----> recombinant DNA molecule

Introduce recDNA into host
Select host and desired recDNA
Propagate recDNA in host, isolate DNA
Bypassing libraries: PCR
Characterize DNA. Expression. Mutagenesis.
Other- special topics

The aim of the lectures is to "flesh out" the above outline, and to give some information necessary to understand the methods used in the lab (some of this should be familiar from BISC 331). We'll then go over the outline in the kind of detail necessary to read the literature, and follow parallel demonstrations of the experiments we'll be doing. Lab exercises will involve one full afternoon per week, plus additional open lab time before and/or after to setup and analyze results. In addition to lecture material listed above, there will be extra material covered in lab handouts and talks.

Accompanying Lab Exercises:

A. Introduction: setting up a lab, safety, equipment, microbiology and sterile technique
B. Preparation and characterization of genomic DNA
C. Plasmid miniprep, restriction digestion and gel electrophoresis
D. Southern blotting
E. Screening a phage library: plaque lifts (Non-radioactive detection of positives etc.)
F. Isolation of DNA and subcloning into a plasmid
G. Polymerase Chain reaction
H. DNA sequencing, dideoxy method, using radioactivity (33P)

TEXT:

Required: C. Howe, Gene Cloning and Manipulation. This will be supplemented by other handouts and materials.

GRADING:

Grading will be based on a midterm (20%), short lab reports plus one complete lab writeup (probably about 20%), a library research project (10%) and a final exam (roughly 50%), exact proportions TBA.