The depth of the mixed layer is a major determinant of nutrient and light availability for phytoplankton in stratified waterbodies. Ongoing climate change influences surface waters through meteorological forcing, which modifies the physical structure of fresh waters including the mixed layer, but effects on phytoplankton biomass are poorly known. To determine the responses of phytoplankton biomass to the depth of the mixed layer, light availability and associated meteorological forcing, we followed daily changes in weather and water column properties in a boreal lake over the first half of a summer stratification period. Phytoplankton biomass increased with the deepening of the mixed layer associated with high wind speeds and low air temperature relative to the temperature of the mixed layer (Tair−Tmix < 0), whereas heatwave conditions—shallow mixed layer driven by high Tair−Tmix value and low wind speed—reduced the biomass. Improving light availability from low to moderate light conditions increased the phytoplankton biomass, while the highest light availability was associated with low phytoplankton biomass. Our study demonstrates that the climatic impact-drivers wind speed and Tair−Tmix are major drivers of mixed layer depth, which controlled phytoplankton biomass during the early summer stratification period. Our study suggests that increasing air temperature relative to water temperature and declining wind speeds have potential to lead to reduced phytoplankton biomass due to a shallower mixed layer during the first half of the stratification period in non-eutrophic lakes with sufficient light availability.