Teaching molecular genetics using <i>Paramecium</i> and RNA interference: research-based learning and project ownership

Research-based course design is beneficial to both the instructor and the students by providing project ownership, independence, increased engagement, and publishable results. <i>Paramecium</i>, a single-celled eukaryote, is a common organism observed in many high school and college classrooms that can be easily cultured and manipulated to navigate through guided student-driven research projects. Presented here are research-centered student projects that include designing and creating an RNA interference (RNAi) plasmid to deplete a gene product in <i>Paramecium</i>. Because RNAi can be used in a large number of model organisms, the techniques presented can be applied in a variety of ways. Using <i>Paramecium</i>, this advanced genetics class uses control and depleted cells to observe changes in cell morphology, cell swimming behavior, and changes in RNA transcript levels. Here, we will describe the use of database searches, primer and construct design, plasmid generation, subcloning, and bacterial screening to generate an RNAi construct and deplete targeted transcript levels. Student data showing the depletion of potential IFT38/40, IFT140, and KATNIP gene products in <i>Paramecium</i> are shared, and these depleted cells show significantly slower swimming speeds with no noticeable change in cell morphology. Overall, students are engaged, invested in their results, and successfully work as collaborative pairs to produce publishable results using this ciliated protist, all while learning cutting-edge molecular techniques.