For more than 20 years, the Gene Transcription in Yeast meeting series biannually brings together researchers using yeast to study mechanisms of gene expression. Owing to the exceptional experimental accessibility of yeasts - in vivo and in vitro, in S. cerevisiae, S. pombe and other yeasts - the yeast community keeps generating both pioneering method development as well as new concepts that often inspired analogous discoveries in the metazoan/human system. The meeting traditionally covers all steps from chromatin to transcription to RNA processing, and integrates different approaches such as structural biology, biochemistry, genetics, cell biology, high-throughput, computational, systems and single-cell technologies. The subtitle “From single molecules to phase separation” reflects both a renewed attention to the individual molecule as well as the will to embrace the exciting discoveries on liquid-liquid phase separation (LLPS) mediating the compartmentalization of many cellular, including nuclear functions.