© 2019 Elsevier B.V. Polysynthetically twinned (PST) crystals of TiAl alloys have attracted much attention due to its unique lamellar microstructure. Their microstructural evolution is essential for understanding the relationship between the microstructure and mechanical performance. Here, we report a comprehensive study of the microstructure and crystallography in Ti-47.5Al PST crystals via a combination of electron backscatter diffraction, in-situ neutron diffraction and high-temperature laser-scanning confocal microscopy during heating cycles. It was found that Σ3<111> twins of γ-phase are unlikely to form along the lath direction. The observable γ crystal orientation variants along the lath belong to the same twin family and are related to each other by a 120° rotation along the <111> direction. Further diffraction studies show that a short-range-ordered structure persists in the disordered α-phase matrix at high temperature and recrystallizes to form a poly-crystalline structure of fully lamellar colonies after heat treatment. The microstructural evolution during heating and cooling is revealed by in-situ high-temperature laser-scanning confocal microscopy, showing the detailed structural transformation and phase evolution from single grain PST crystals into a poly-crystalline structure.