Abstract: The technique of pollen-mediated plant genetic transformation assisted by ultrasonication was patented by the Biotechnology Research Center of the Shanxi Academy of Agricultural Sciences. Here, we further investigated the conditions for sampling, storage and treatment of maize (Zea mays L.) pollen and effects on in vitro germination of pollen with the method. The optimal sucrose concentration for pollen suspension solution differed for maize pollen collected from plants with different flowering time. We determined the storage time and condition of pollen and their effects on pollen germination after ultrasonication. These results would be useful for improving pollen vitality in genetic transformation and verifying the feasibility of this plant transformation technique. We discuss some key steps for conducting in vitro germination of maize pollen and related parameters to increase transformation efficiency of pollen-mediated plant genetic transformation.