Petri nets are a useful mathematical formalism for specification of manufacturing systems, supported by various analysis and verification methods. The progress made in automating control systems and the widespread use of Industry 4.0 pose a number of challenges to their application, starting from the education at university level and ending with modelling of real case studies. The paper aims to present and analyse the most relevant challenges and opportunities related to the use of Petri nets as a modelling technique of manufacturing systems. The review of the literature is primarily based on the years 2019–2020 to reflect the current state of the art. The newest approaches to deadlock prevention and recovering, but also other important analysis problems and difficulties in modelling real industrial processes are discussed. Trends for the future are also identified.