A Versatile Semester-Long Course-Based Undergraduate Research Experience using Optogenetics and RNAi to Identify Genes Important for Synapse Function

Abstract

Compared to traditional teaching laboratory activities, course-based undergraduate research experiences (CUREs) can increase student engagement and confidence, improve scientific literacy, enhance critical thinking, and promote accessibility in STEM. Here we describe a versatile CURE for an upper-level Neurobiology course that incorporates genetic, molecular, cellular, and behavioral experiments into a semester-long investigation to identify genes important for glutamate synapse formation or function in C. elegans. Following introduction to the CURE approach and basic C. elegans techniques, students construct their own low-cost optogenetics rigs, which we describe in detail here, to activate a mechanosensory escape reflex via photostimulation. They then perform a small-scale RNAi screen with this light-activated behavioral readout. Once a gene of interest is identified, students submit a proposal to investigate the role of this gene in nervous system function and spend the rest of the semester carrying out follow-up experiments using mutant strains. We also describe ways in which this CURE can be modified depending on the pedagogical objectives, availability of materials, or research interests of the instructor. Participating in this lab significantly enhanced students' abilities to see themselves as STEM professionals and prompted students to report substantial gains in skills critical for entry into and success in graduate and medical schools. In addition to the benefits CUREs provide to students, faculty benefit from the generation of preliminary data and training of students for potential independent research projects.

Keywords: (CURE); Caenorhabditis elegans; Course-Based Undergraduate Research Experience; RNAi; behavior; low-cost; optogenetics.

Figure 1

RNAi screen overview. Egg-bearing worms are added to standard worm growth plates containing RNAi bacteria and all-trans retinal (ATR), a necessary cofactor for the light-responsive channelrhodopsin (ChR2) protein. One week later, offspring with genes knocked down are scored for glutamate-independent gross locomotor activity (thrashing) and glutamate-dependent reflex behavior, which is triggered by blue light activation of ChR2 in sensory neurons. Based on the results of these behavioral tests, students identify and investigate a candidate gene for which a mutant strain is then ordered from the Caenorhabditis Genetics Center.

Figure 2

Assembly of the LED illuminator. A) LED components: 1. LED light kit 2. LED driver 3. Power adapter 4. Wall mount switch 5. Foot switch. Boxed regions are represented in B and C. Red arrows indicate additional connection sites. B) Red and black wires from the LED driver should be connected to the + and − terminals, respectively, of the power adapter. C) LED driver and light kit should be wired so that black is connected to blue and red to white.

Figure 3

Measuring power output of a student-assembled LED illuminator. Assembled LED rig is positioned approximately 10 cm above the stage of a dissecting microscope. Light intensity at the point of observation is measured using a silicon photodiode sensor connected to a digital power and energy console.

Figure 4

Student STEM Identity scores before and after the CURE. In response to a multiple-choice item, students selected the option illustrating how much their identity overlaps with their idea of a “STEM professional”. * p = 0.02 Paired T-test