Bacterial Diversity Studies Using the 16S rRna Gene Provide a Powerful Research-Based Curriculum for Molecular Biology Laboratory

S. Boomer, D. Lodge, and B. Dutton (Western Oregon University)

Abstract

Over the past three years, we have developed and implemented a ten-week curriculum for our molecular biology course based on our microbiology research project. Our project is centered around the isolation and characterization of 16S rRNA genes from hot spring bacterial communities in Yellowstone National Park. Data generated in this course consistently demonstrate novel and diverse bacterial sequences, expanding our collective understanding of microbial diversity and allowing students to learn about microbiology while mastering a complex series of molecular and computational methods. For the first five weeks of the course, students prepare plasmids from previously isolated 16S rRNA clones and characterize each via restriction mapping and nucleotide sequence analysis. Next, they isolate genomic DNA from microbial mats, utilize PCR to amplify 16S rRNA genes, and clone products. Each student screens a portion of his/her library, mastering small-scale plasmid preparation and re-applying restriction mapping methodology to assess for the presence of 16S rRNA insert. Depending on time and progress, students may determine the nucleotide sequences of insert-bearing clones. During the final two weeks of the course, students classify and infer identity of all obtained 16S rRNA isolates using computer-based phylogenetic methods and databases.

This overall order of topics was chosen because techniques used to characterize clones are more forgiving than sensitive cloning and PCR methods. As the course is offered each year, it naturally accumulates new clones for subsequent years. We contend, given the ubiquity and diversity of microorganisms, that this research-based curriculum can be adapted to a variety of sample sources to elucidate microbial diversity that relates to medical/health, applied, or environmental microbiology. Finally, the approach advocated here is ideal for exposing pre-service education majors and post-baccalaureate teachers to inquiry-based science curriculum strategies.