This book constitutes the refereed proceedings of the 20th International Symposium on Bioinformatics Research and Applications, ISBRA 2024, held in Kunming, China, in July 19–21, 2024. The 93 full papers included in this book were carefully reviewed and selected from 236 submissions. The symposium provides a forum for the exchange of ideas and results among researchers, developers, and practitioners working on all aspects of bioinformatics and computational biology and their applications.