A course-based undergraduate research experience (CURE) using transposon-directed insertion-site sequencing (TraDIS)...

Understanding gene function through forward genetic screens is foundational to microbial genetics. Here, we describe the design and implementation of a multi-week course-based undergraduate research experience (CURE) embedded within a third-year Microbial Genetics course, developed over three iterations in trimester 3 of 2023-2025. In this CURE, students construct a high-density Tn5 transposon insertion library in Escherichia coli, challenge the library with bacteriophage P1, and use transposon-directed insertion-site sequencing (TraDIS) coupled with statistical analysis (edgeR) to identify genes required for phage propagation. Students performed transposome electroporation, phage infection assays, genomic DNA extraction, analysis of high-throughput sequencing data on Galaxy, differential insertion analysis, data visualization, and ontology enrichment analysis. The project integrates wet-lab microbiology, molecular genetics, and bioinformatics while modeling an authentic forward genetic screen. This CURE emphasizes experimental design, genome-wide functional analysis, statistical reasoning, and interpretation of large-scale sequencing data. The curriculum is adaptable to other bacterial species, phage systems, or selective pressures, and provides a scalable framework for integrating authentic genomics research into upper-level microbiology courses.