Ficedula flycatchers are important avian speciation models. In particular, the naturally hybridizing collared flycatcher (Ficedula albicollis) and pied flycatcher (Ficedula hypoleuca) have been intensively studied in the context of speciation. Collared flycatchers and pied flycatchers are two closely related migratory passerine birds from the family Muscicapidae, which diverged approximately 1 MYA. They winter in sub-Saharan Africa and breed in woodlands across Europe, where they have several areas of sympatry and occasionally hybridize. Their F1 hybrids suffer from low to totally compromised fertility, low attractivity, and abnormal metabolic rate, which makes them a relevant model system for speciation research. A rich body of ecological information, a chromosome level genome assembly of the collared flycatcher, and protein-coding sequence, repeat sequence as well as conserved non-coding elements annotations have been established and are publicly available. Together with collaborators, we decided to generate high-quality genome assemblies of both species with help of PacBio long-read sequencing data in combination with Hi-C data, as well as to improve the genome annotation with help of PacBio Iso-Seq data. This data resource shall enable us to establish a more complete genome assembly and contribute to the identification of ‘missing genes’ in avian genomes. Moreover, the generation of population-level PacBio long-read sequencing data for both species puts us in the position to study the role of structural variants in speciation. Second, besides evolution at the sequence level, we also investigate changes at the gene expression and epigenetic level among collared flycatchers, pied flycatchers and their F1 hybrids. This second research angle helps to improve our understanding of the functional importance of different genomic features and their interaction with each other in the hybrid genomic background. Third, we recently generated population-level short-read sequencing data across 10 distinct populations of pied flycatchers, in order to characterize build-up of reproductive isolation at shorter time-scale. For this third research angle we will combine genomic data with extensive sampling of songs across populations to understand the speed and scale at which song divergence emerges and how it is linked with reproductive barriers across an entire species range. The total project thus contains of three smaller projects, ‘The role of structural variants in speciation of collared flycatchers and pied flycatchers’, ‘Molecular mechanisms of hybrid incompatibilities among collared flycatchers and pied flycatchers’ and ‘The genomic basis of song divergence in pied flycatchers’. We propose to store sequencing data in a medium storage project and perform bioinformatics pipelines in an associated compute project.