Temperature as a determinant factor for increased and reproducible in vitro pollen germination in Arabidopsis thaliana

Despite much effort, a robust protocol for in vitro germination of Arabidopsis thaliana pollen has been elusive. Here we show that controlled temperatures, a largely disregarded factor in previous studies, and a simple optimized medium, solidified or liquid, yielded pollen germination rates above 80% and pollen tube lengths of hundreds of microns, with both Columbia and Landsberg erecta (Ler) ecotypes. We found that pollen germination and tube growth were dependent on pollen density in both liquid and solid medium. Pollen germination rates were not substantially affected by flower or plant age. The quartet1 mutation negatively affected pollen germination, especially in the Ler ecotype. This protocol will facilitate functional analyses of insertional mutants affecting male gametophyte function, and should allow detailed gene expression analyses during pollen tube growth. Arabidopsis thaliana can now be included on the list of plant species that are suitable models for physiological studies of pollen tube elongation and tip growth.