Next Generation Sequencing (NGS) has become an important tool in the biological sciences and has a growing number of applications across medical fields. Currently, few undergraduate programs provide training in the design and implementation of NGS applications. Here, we describe an inquiry-based laboratory exercise for a college-level molecular biology laboratory course that uses real-time MinION deep sequencing and bioinformatics to investigate characteristic genetic variants found in cancer cell-lines. The overall goal for students was to identify non-small cell lung cancer (NSCLC) cell-lines based on their unique genomic profiles. The units described in this laboratory highlight core principles in multiplex PCR primer design, real-time deep sequencing, and bioinformatics analysis for genetic variants. We found that the MinION device is an appropriate, feasible tool that provides a comprehensive, hands-on NGS experience for undergraduates. Student evaluations demonstrated increased confidence in using molecular techniques and enhanced understanding of NGS concepts. Overall, this exercise provides a pedagogical tool for incorporating NGS approaches in the teaching laboratory as way of enhancing students' comprehension of genomic sequence analysis. Further, this NGS lab module can easily be added to a variety of lab-based courses to help undergraduate students learn current DNA sequencing methods with limited effort and cost.