In the Mediterranean Sea near Marseille, Italian fishermen rescue an unconscious American man floating adrift with two gunshot wounds in his back. The boat's medic finds a tiny laser projector surgically implanted under the man's skin at the hip. When activated, it displays a safe deposit box number in Zürich. The man wakes up and discovers he is suffering from memory loss. Though fluent in several languages and possessing unusual skills, he cannot remember anything about his identity or events prior to his rescue. When the ship docks, he sets off to investigate the deposit box.
At CIA headquarters in Langley, Virginia, Deputy Director Ward Abbott learns that a CIA assassination attempt against exiled Nigerian dictator Nykwana Wombosi failed. Meanwhile in Zürich, the amnesiac man incapacitates two policemen using advanced hand-to-hand combat when they attempt to arrest him. The next morning, he visits a bank and opens the safe deposit box of the account number displayed by the implanted projector to find several passports containing his picture under different names, nationalities and addresses, large amounts of assorted currencies, and a handgun. Taking an American passport name, Jason Bourne, he leaves with everything but the handgun.
As he leaves, a bank employee contacts Operation Treadstone, a CIA black ops program. Alexander Conklin, head of Treadstone, assures Abbott that he will clean up Bourne's failure against Wombosi, activating three agents to take down Bourne: Castel, Manheim, and The Professor. Chased by police, Bourne is then pursued through a U.S. consulate. He encounters a German woman named Marie Kreutz, offering her $20,000 to drive him to the address in Paris on his French driving licence.
Bourne arrives at the address, an apartment, where he hits redial on his phone and is connected to Hotel Regina. Bourne enquires about some names: a "Jason Bourne" is not registered there, but "John Michael Kane" was and died two weeks ago in a car crash. Castel then ambushes Bourne and Marie at the apartment, but Bourne defeats him. Marie finds wanted posters in Castel's bag with both her and Bourne's pictures, while Castel jumps out of a window to his death. Marie, in shock, is later resolved to join Bourne as they escape and investigate "John Michael Kane".
Wombosi had contacted police about the attempt on his life. To convince him that his assailant, Kane, is dead, Conklin plants a body in a Paris morgue but Wombosi is not fooled. The Professor then assassinates Wombosi as a result. Bourne poses as Kane and gains documents about Wombosi's yacht and finds that an assailant was shot twice while escaping it. He concludes that he is the assassin, before seeking refuge in the French countryside with Marie and her step-brother, Eamon and his children. Conklin traces their location and The Professor arrives in the morning. Bourne grabs Eamon's shotgun and outflanks The Professor in the fields, mortally wounding him. The Professor reveals their mutual connection to Treadstone before dying. Bourne sends Marie away with Eamon. He contacts Conklin using The Professor's phone to arrange a meet, but Conklin brings back-up, so Bourne instead tracks his vehicle to locate Treadstone's Paris safe house.
Bourne breaks in and holds Conklin and logistics technician Nicky Parsons at gunpoint, beginning to remember his last mission through successive flashbacks. Kane was Bourne's assumed identity during his mission to infiltrate Wombosi's yacht and kill him, but Bourne failed to because Wombosi's children were sleeping nearby. Bourne fled and was shot twice in the back in the process. Bourne tells Conklin that he is leaving Treadstone and warns not to follow him. After their altercation, both Conklin and Bourne leave, with Bourne fighting his way out through more agents. A nearby Manheim steps out of his car and kills Conklin: Abbott, in order to terminate Treadstone, ordered the hit. Before an oversight committee, Abbot dismisses Treadstone and announces a new project codenamed "Blackbriar". Some time later, Jason finds Marie renting out scooters to tourists in Mykonos, Greece and the two reunite.