Understanding how phytoplankton adapt to elevated CO₂ and/or warming through long-term genotypic changes is critical for predicting future phytoplankton distribution and community structure. In this study, we conducted a 4.5-year experimental evolution with the model marine diatom Phaeodactylum tricornutum Bohlin under four environmental conditions: ambient (control), elevated CO₂, warming and combined elevated CO₂ + warming. Following this long-term adaptation, we exposed the populations to a broad CO₂ gradient in a short-term (7-day) experiment to assess their multi-trait responses. Our results demonstrate that P. tricornutum Bohlin populations adapted to different environmental regimes exhibit significant multi-trait variation across CO₂ gradients. Notably, the variability driven by long-term adaptation exceeded that induced by short-term CO₂ changes. Furthermore, both long-term adaptation and short-term CO₂ exposure altered trait co-variations, highlighting the complex interplay between environmental history and immediate conditions. This study emphasizes the importance of assessing long-term genetic changes in marine phytoplankton under global change, as short-term experiments alone may underestimate their adaptive potential and the broader implications for marine ecosystems under future climate scenarios.