Microalgae can be used for biodiesel production, but not all algae can grow well in different environments. Parachlorella kessleri TY was grown with glucose in different ways, and mixotrophic cultivation led to the highest biomass and lipid accumulation. Both heterotrophic and mixotrophic conditions increased lipid content, especially neutral lipids and fatty acids suitable for biodiesel. Mixotrophic cultivation is better for lipid accumulation in P. kessleri TY.