Lincoln Six Echo (Ewan McGregor) lives in an isolated compound which strictly regulates its inhabitants' lives. The Overseers control every aspect of the residents' lives, from diet and free time activities to social relationships. The inhabitants hope to win a lottery to go to "the Island", the only place on Earth not contaminated by a deadly pathogen. Lincoln tells Dr. Merrick (Sean Bean) of his discontent with the plain white clothes everyone is given and the strict social controls. Concerned about strange dreams Lincoln experiences about a speedboat and other unfamiliar items, Merrick inserts probes into Lincoln's body to monitor his cerebral activities.
While illicitly visiting a power-plant basement where his friend, the technician James McCord (Steve Buscemi), works, Lincoln discovers a live moth in a ventilation shaft, leading him to deduce that the outside world is not contaminated. Lincoln captures and releases the moth and follows it to another section, where he witnesses two recent lottery winners being killed: one after childbirth, and the other in the process of having his liver harvested. Security discovers that Lincoln has wandered into unauthorized areas, and starts searching for him. As Lincoln rescues Jordan Two Delta (Scarlett Johansson), a friend who has just won the lottery, Dr. Merrick discovers that Lincoln is beginning to discover the truth.
While eluding security and escaping from the facility, Lincoln and Jordan learn that both the contamination of the outside world and the Island are myths. And later learn that the residents are clones of wealthy sponsors, who intend to use them for spare parts or surrogate motherhood. Merrick's technology allows clones to incubate directly into adulthood. He tells sponsors and the government that the clones do not gain consciousness, but when Merrick hires French mercenary Albert Laurent (Djimon Hounsou) to kill or capture the escapees, he admits that the organs of unconscious clones inevitably fail, making them useless. Thus, he needs the clones conscious in the compound.
Lincoln and Jordan escape the compound and find themselves in the Arizona desert. They find McCord, who gives them the name of Lincoln's sponsor in Los Angeles, and helps them to the Yucca railway station. After Laurent's mercenaries find and kill McCord, Lincoln and Jordan ride the train in hopes of finding help in Los Angeles. (Jordan Two's sponsor, model Sarah Jordan, cannot help as she is in a coma following a car accident.) Lincoln Six's sponsor, Tom Lincoln—the owner of the speedboat in Lincoln's dreams—agrees to help expose the truth about the organ harvesting, but secretly turns against his clone by informing Merrick about the situation. Merrick sends the mercenaries to the location, but Lincoln Six tricks Laurent into killing Tom and assumes Tom's identity.
The cerebral probes reveal that Lincoln Six Echo inherited some memories from his sponsor Tom Lincoln (which had been assumed impossible). Merrick becomes afraid that the same will happen with the other clones and decides to kill them all. Lincoln and Jordan plan to liberate their fellow clones. Now posing as Tom Lincoln, Lincoln Six returns to the compound in order to destroy the holographic projectors that hide the outside world from its residents. With help from Laurent—who has moral qualms with treating the clones as if they are not human—and Jordan, Merrick is killed and the clones are freed. The film ends with a shot of the clones on hill outdoors, seeing the outside world for the first time, and then the film cuts to the final frame which shows Lincoln and Jordan together on the deck of the speedboat from his dreams.
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