ABSTRACT In the search for life in the Universe, Earth provides a template of evolution for the one habitable planet we know. Earth’s atmospheric composition has changed significantly throughout its history. The last 500 Myr – the Phanerozoic Eon, which includes the origins of animals, dinosaurs, and land plants – saw oxygen rise from ≤10 per cent to 35 per cent. But the resulting transmission spectra are a crucial missing piece in our search for signs of life in exoplanet atmospheres. Here, we simulate the atmosphere and transmission spectra of the Phanerozoic, using estimates from established climate models, and present the first high-resolution transmission spectra for Phanerozoic Earth. We demonstrate that the spectral biosignature pairs O2 + CH4 and O3 + CH4 in the atmosphere of a transiting Earth-like planet would indicate a biosphere, with O2 and O3 features potentially stronger than for modern Earth. The full model and high-resolution transmission spectra, covering 0.4–20 µm, are available online and provides a tool to plan and optimize observations, train retrieval methods, and interpret upcoming observations with ground- and space-based telescopes.