Genomics has revolutionized biological research, but quality assessment of the resulting assembled sequences is complicated and remains mostly limited to technical measures like N50. We propose a measure for quantitative assessment of genome assembly and annotation completeness based on evolutionarily informed expectations of gene content. We implemented the assessment procedure in open-source software, with sets of Benchmarking Universal Single-Copy Orthologs, named BUSCO. Software implemented in Python and datasets available for download from http://busco.ezlab.org. firstname.lastname@example.org Supplementary data are available at Bioinformatics online.