c-Myc Knockout as a Model for Gene Editing for Training Healthcare Professional Students

Abstract

Abstract Correction of genetic errors, commonly known as gene editing, holds promise to treat diseases with unmet medical needs. However, several gene therapy trials have encountered unwanted outcomes, because of the incomplete understanding of the disease states and gene therapy processes. This situation encourages a concept that healthcare professionals receiving laboratory research will not only identify inadequacies in basic biomedical knowledge in gene therapy but also provide tangible refinements. We thus have undertaken the PharmD student-training program in gene editing in a basic research laboratory setting. As a model, MYC gene was chosen for knockout using CRISPR-Cas9 method in HT29 and OVCAR8 cells. Students were involved in the design of MYC-specific gRNAs, subcloning into Cas9-carrying plasmid, and selection of knockout clones from the transfected cells. Students were involved in the genomic DNA isolation, analysis of DNA sequences from the clones using online bioinformatics tools, western blotting, cell proliferation and cell division cycle, to characterize the knockout clones. Observations made during this training are presented. In this process, students have gained a better understanding of the disease mechanisms, gene therapy protocols, limitations of gene therapies, abilities to evaluate the literature and transfer laboratory training to the practice of gene therapies in clinic.