Alkenone‐derived paleotemperature reconstruction holds great promise in lake environments. However, the occurrence of multiple species of alkenone‐producing haptophyte algae in a single lake can complicate the translation of alkenone unsaturation to temperature if each species requires an individual temperature calibration. Here, we present the first systematic monitoring of two alkenone‐producing haptophytes throughout the course of a seasonal cycle in Lake George, North Dakota, using a combined approach of DNA sequencing and alkenone lipid characterization. Field sampling revealed a nonoverlapping haptophyte succession, with both an early and late season haptophyte bloom event. Culturing experiments demonstrated that the two haptophyte species responsible for these blooms had statistically similar alkenone‐temperature responses, although the culture‐based calibrations were distinct from the in situ calibration. Bloom timing of each haptophyte species corresponded to surface‐water temperatures that differed by more than 10°C, revealing that changes in bloom intensities for each species will skew the sediment‐inferred temperatures to a different stage of the growth season. These results highlight the importance of accounting for bloom timing when interpreting alkenone‐derived temperatures in sediment cores, especially in lakes that experience large seasonal fluctuations in water column temperature and salinity.