Pseudomonas aeruginosa, a typical opportunistic pathogen, has a single polar flagella which provides swimming motility and virulence. Twelve of insertional mutants with Mu transposition complexes into genomic DNA of Pseudomonas aeruginosa were screened on the changes of swimming motility. After gene cloning and DNA sequencing of those mutants, ten of them were illustrated that defective mutations on swimming motility were caused by insertions of Mu transposon into 10 of different functional genes which may involved in biosynthesis of flagella and gene regulations, but another two mutants were resulted from the insertion of two new genes (PA2950, PA5022) with function unknown. Observation of these two mutants with an integrity flagellum under electron microscope showed that two new genes were not involved in construction of flagella, but probably in energy metabolism and gene regulation of swimming motility.