Background: Pharmacogenomics (PG) is a modern tool of personalizing treatment protocols to improve the efficacy and safety of drug prescriptions. These benefits are offset by a slow uptake in clinical application due to a host of physician factors, patient factors, and/or health system factors. Our study, thus, aimed to determine the knowledge, attitude, future expectations, and perceived barriers of medical students and physicians in Jordan regarding PG testing. Methods: A descriptive, cross-sectional study was conducted between February-August 2019. Physicians and senior medical students from academic and non-academic institutions in North Jordan (n=424) were surveyed. A structured, self-administered questionnaire was designed and piloted for the purpose of the study. A scoring system for each dimension assessed was calculated and presented using means. Mean scores were compared by sociodemographic and professional variables. Results: The response rate was 70.7%. The mean total PG knowledge score (±SD) was 5.42 (±1.51) out of 10, with a significantly higher mean among respondents aged ≥30 years (5.21 ± 1.62) compared to those <30 years-old (5.54 ± 1.43; p= 0.03). The mean total PG attitude score was 21.18 (±2.58) out of 24, with significant differences by seniority levels evident (p= 0.03). The future expectations of PG among our sample were high, with a mean score of 10.44 (±1.64) out of 12. The top three perceived barriers in applying PG were the high cost, lack of clinical guidelines, and limited knowledge and awareness. Conclusion: Physicians and medical students in Jordan have low overall knowledge, albeit strongly positive attitude and future expectations toward PG, despite the perceived high cost and lack of clinical guidelines. Thus, we strongly recommend adopting a comprehensive educational strategy that aims to integrate PG concepts into medical curricula, and promote the culture of continuous medical education about PG among practitioners.