Background <p>The aim of this study was to identify and prioritise the ten most important unanswered themes in rare disease research in Australia by integrating perspectives of key stakeholders, including people living with rare disease, parents/carers, health professionals, and rare disease community advocates.</p> Methods <p>We conducted a Priority Setting Partnership project based on a modified James Lind Alliance Priority Setting Partnership methodology. The process involved an online elicitation survey, review of existing literature, and consensus workshops with Australian rare disease stakeholders.</p> Results <p>In the elicitation survey (<i>n</i> = 185), 585 questions/comments were coded, and 19 overarching themes identified. No themes were completely addressed by existing literature. In the consensus workshops (<i>n</i> = 34), the 19 themes were refined, and the Top 10 priorities determined. The top three priorities were (1) development of, and access to, the best treatments and cures, (2) awareness and education for health professionals and service providers, and (3) diagnosis, including screening.</p> Conclusion <p>The Top 10 priorities span a wide range of health research domains, including access to effective diagnosis and treatment, healthcare system capacity and expertise, and support for patients, families, and carers.</p> Patient or public contribution <p>A Project Advisory Group and Steering Committee were established to guide this study. The Project Advisory Group met four times across the duration of the project, while the Steering Committee met three times to provide feedback and support recruitment and translation of findings. </p>