Development of a Freeze-Dried CRISPR-Cas12 Sensor for Detecting Wolbachia in the Secondary Science Classroom

Training the future synthetic biology workforce requires opportunity and exposure to biotechnology concepts and activities in secondary education. Detecting Wolbachia bacteria in arthropods using PCR has become a common way for secondary students to investigate and apply DNA technology in the science classroom. Despite this framework, cutting-edge biotechnologies like CRISPR-based diagnostics have yet to be widely implemented in the classroom. To address this gap, we present a freeze-dried CRISPR-Cas12 sensing reaction to complement traditional DNA technology education and teach synthetic biology concepts. The reactions accurately detect Wolbachia from arthropod-derived PCR samples in under 2 hours and can be stored at room temperature for over a month without appreciable degradation. The reactions are easy-to-use and cost less than $40 to implement for a classroom of 22 students including the cost of reusable equipment. We see this technology as an accessible way to incorporate synthetic biology education into existing biology curriculum, which will expand biology educational opportunities in science, technology, engineering, and mathematics (STEM) education. ### Competing Interest Statement M.C.J. is a cofounder of SwiftScale Biologics, Stemloop, Inc., Design Pharmaceuticals, and Pearl Bio. M.C.J. interests are reviewed and managed by Northwestern University in accordance with their conflict of interest policies. All other authors declare no competing interests. * PCR : Polymerase Chain Reaction CRISPR : Clustered Regularly Interspaced Short Palindromic Repeats Cas : CRISPR Associated Protein FD-CC12 : Freeze-Dried CRISPR-Cas12 STEM : Science Technology Engineering and Mathematics K-12 : Kindergarten through Twelfth Grade ssDNA : Single Stranded DNA FQ : Fluorophore Quencher PAM : Protospacer Adjacent Motif cr crRNA : CRISPR RNA