200 years after the events of Alien 3, scientists aboard a spaceship called the USN Auriga are working on a secret project for the military; they have been attempting to produce a clone of Ellen Ripley (Sigourney Weaver) working from a blood sample that was collected from Fiorina 161, the planet where she died. In doing so, they hope that they can harvest the alien embryo, a queen of the species, that was gestating inside of her at the time. Dr. Wren (J.E. Freeman) and his team of colleagues have succeeded with their eighth attempt. They operate on the Ripley clone and remove the embryonic alien, which is still too small to struggle with the robotic arms used for the operation. After the alien is removed, Dr. Wren and his colleague, Dr. Gediman (Brad Dourif), decide to keep the clone alive for further study. They repair the wound in her chest. As she recovers, she notices a numeral 8 tattooed on one of her arms.
The Ripley clone begins to gain consciousness and mature at an unprecedented rate. The ship's doctors go through exercises with her regarding language and learning, and she responds positively. She also exhibits a frightening predatory physical power, able to break her bonds and attack at will. The doctors and orderlies are only barely able to subdue her. She seems to share DNA with the alien creatures, explaining her unusual strength, and her blood is also slightly acidic. The experiment angers General Perez (Dan Hedaya), the military figure in charge of the operation, who calls her a "meat byproduct". He's only interested in the alien queen, which has also grown at a terrifying rate; it is now almost full size and is contained within a cell on the ship.
The Ripley clone seems to sense what's going on. One day, she eats with Gediman and Wren, and she reveals a startling flash of her former self; it seems as if the clone has retained some of Ripley's memories and personality traits. It is almost as if the woman herself has been recreated and is slowly awakening. Ripley has a fragmented conversation with Gediman. "How did you...?" she asks. Gediman tells her how they painstaking recreated her, and she intuitively guesses that they wanted the alien. Even in her dazed condition, she is cynical and tells them that they can't control it.
Enter the Betty, a ragtag ship carrying a crew of space pirates. Their cargo is the crew of another ship, still in their cryogenic containers and asleep, unaware that they have been hijacked. General Perez has hired the crew, led by Frank Elgyn (Michael Wincott), to deliver the unfortunate sleeping travelers to be used for breeding the aliens. Before long the kidnapped people have been impregnated with aliens, and a number of drone aliens are now in containment units on board the ship. The Betty crew find Ripley exercising in a recreation center, tossing a basketball around. Johner (Ron Perlman), one of the Betty's crew and a brutish, scarred man, attempts to engage her in a predatory manner of flirting, but Ripley easily outmaneuvers and fights back against him and one of his comrades, Christie (Gary Dourdan), and proves herself to be physically powerful beyond their expectations. At one point, her nose is bloodied; when she wipes off the blood and flicks it onto the floor, it begins to dissolve the surface, much like the blood of the aliens.
One of the Betty's crew, a young waifish girl named Annalee Call (Winona Ryder), seems to have a different agenda. She slips away from the others and finds Ripley's containment unit, gaining illegal access to it. There she finds Ripley and wakes her up; Call admits that she came here to kill Ripley, but Ripley is far too cunning to allow that. She even deliberately presses her hand onto Call's knife; the resulting wound heals itself rapidly and her blood corrodes the blade. Call can tell that the doctors have removed the alien embryo already, and she tells Ripley that she is just a clone, something the scientists consider only a byproduct. Ripley seems confused and hurt by this, but she knows it is true. She releases Call, who goes outside the cell only to be intercepted by guards and Dr. Wren.
Wren is furious about Call's trespassing, and he attempts to execute the crew of the Betty as terrorists. A number of things happen simultaneously. The Betty crew has a battle with soldiers who have orders to execute them. While experimenting with adult aliens in a cell, Gediman inadvertently teaches them how to use the pacifying equipment in the cell, which sprays freezing gas. Together the aliens revolt; two of them gang up on a third and kill it, causing its acid blood to eat through the floor of the containment unit. Gediman is snatched away by one of the aliens. Now loose, the aliens quickly begin to take over the ship. Ripley escapes from her cell when aliens begin to break through the door. The ship goes into emergency mode and some people escape in lifepods; the unfortunate ones are killed by aliens. The crew of the Betty bands together, but Elgyn is killed by the creatures.
Ripley joins Call and the others as they make their way across the Auriga to the bay where the Betty is docked. Ripley also discovers that the Auriga is actually moving. Wren scoffs, saying she couldn't possibly know because the ship is equipped with stealth run. However, Ripley's heightened senses are correct, the ship is moving, traveling rapidly back to homebase: Earth. The crew of the Betty is disheartened since Earth has become a polluted wasteland. They take Wren with them, although he is more of a hostage.
Along the way, they find the laboratory where Gediman and the others created the Ripley clones; inside are the previous seven attempts, all of them hideously deformed and bizarre human/alien hybrids. Six are dead and contained in stasis tubes, but the seventh attempt is the most heartbreaking; she is human enough to be identifiable, yet her limbs and her body are misshapen and alien. Even worse, she is still living, kept alive by tubes and ventilators. Ripley 8 is horrified, realizing now exactly what she is and where she came from. Ripley 7 begs her to kill her, and Ripley 8 uses a flamethrower and grenades to destroy the other clones. Wren is morbidly amused by her inner turmoil over her bizarre identity, and Ripley 8 looks at him in utter rage and disgust. Johner has no idea what caused Ripley's rampage; "Must be a chick thing!" he quips.
The crew makes another strange discovery: the room containing what's left of the hijacked crew that the Betty crew delivered to General Perez. One of the crew members, Purvis (Leland Orser), is still alive, but Ripley suggests they leave him behind. She can sense that he is gestating an alien embryo, having been impregnated by a facehugger. Call insists that they bring him along, that perhaps if they make it to the Betty in time, they can put him in cryostasis long enough for some sort of surgery to be performed on him. A hitch in their escape plan occurs when they find that they are forced to traverse the mess hall; unfortunately, it is completely underwater now due to a flood that occurred during the chaos of the alien escape. Vriess (Dominique Pinon) has to leave his powered wheelchair behind, and is harnessed to Christie's back. Everyone must hold their breath for a painful amount of time, and Sabra Hillard (Kim Flowers) from the Betty is snatched along the way by aliens that emerge, swimming after her. Ripley watches, strangely fascinated by the creatures.
When they emerge on the other side of the tunnel, they find that they have been ambushed; the shaft leading upwards from the mess hall has been festooned with alien eggs, which eject facehuggers at the humans. Ripley gets one on her face, but her inhuman strength allows her to rip it away from herself (a strange twist on the similar incident that happened to her in Aliens). Grenades clear the shaft of eggs and allows the crew to climb the ladder to the top, but aliens emerge from the water below them and attack. When Call and Wren reach the top of the shaft first, Wren talks her into giving him her gun and shoots her in the chest; her body falls back down into the water and drifts away. Ripley is horrified, as she has found something strangely compelling about Call. Christie is wounded and grabbed by an alien, and then sacrifices his life to give Vriess a chance to get away. Wren has escaped the shaft and sealed everyone else inside, but the door eventually opens again from the other side; Call has somehow survived the gunshot and rescued them.
Ripley is not deceived, as she knows she saw Call get shot in the chest at close range. She makes Call show everyone the wound, revealing wires and white android blood. Call confesses that she is one of a legendary second generation of androids who were so lifelike that they gained consciousness and rebelled, burning their internal wireless modems and trying to pass themselves as human. The soldier Vincent Distephano (Raymond Cruz) who is with them is geeked about actually seeing this kind of android; Call seems humiliated that her secret has been discovered. Ripley now understands why she feels a kinship with Call; both of them are somehow artificial, creatures that have been created in the likeness of human beings but who are distinctively different.
Knowing Call's capabilities, Ripley takes her to a small chapel, where they find a computer interface. Although Call does not want to make use of her android functions, Ripley urges her to take over the Auriga's computer. Call discovers that the Auriga went into emergency mode after the attack; it is now automatically returning to Earth, where surely the aliens will escape and infect the human race. The ship has burned too much energy for Call to make critical mass and blow it, so Ripley tells her to crash the ship. Call blocks Wren from reaching the Betty and makes a sarcastic announcement summoning all aliens to the deck where he is. Call reveals that before the recall, she accessed the mainframe and discovered the plans that General Perez had for cultivating the aliens by cloning Ripley. Her personal mission was to destroy the operation and keep this from happening, as she understands that the aliens will wipe out humanity.
When they set out for the Betty again, Ripley is snatched through the floor grating by aliens. Her connection to the creatures comes full circle; they recognize her as one of their own and take her to the nest, which is crawling with aliens and which also contains the queen. Gediman is there as well, cocooned and delirious, and he tells Ripley that the queen inherited Ripley's human reproductive cycle. After laying her first round of eggs, she gives birth as a human being would, developing a strange, uterus-like pod and birthing a bizarre creature that resembles a human being crossed with an alien. The Newborn kills the queen and embraces Ripley as its mother. It attacks Gediman, biting off the top of his head. Ripley escapes the nest and races to rejoin the others as the Betty takes off. She makes it just in time, making an impossibly long leap from the deck to the ship. With their pilots dead, Ripley seems to be the only one of them who has the experience to pilot the ship. "Are you kidding?" she tells them, "This piece of shit's older than I am."
There is final confrontation when Call goes to close the bay door and discovers the Newborn has stowed away. She barely manages to evade it. Distephano comes into the bay and is killed. Ripley senses that something is wrong and comes back to find the Newborn cornering Call. She distracts it by speaking soothingly to it; it trusts Ripley as it thinks she is its mother. She deliberately cuts her hand on its sharp teeth and flicks her acidic blood onto a glass porthole. Her blood dissolves a hole in the glass, depressurizing the cabin and sucking the Newborn onto the window. It hovers there pitifully as it is slowly drawn violently out through the tiny hole; Ripley feels torn between her desire to exterminate the species and the loyalty she feels toward them since she has their DNA. She watches in horror as the Newborn is slowly destroyed, aware that it is dying.
The Auriga crashes on Earth and is destroyed. The Betty breaches Earth's atmosphere safely and Ripley and Call look out the porthole at the clouds and the sun, dazed by their experience and wondering what the future will hold for both of them. Neither one has ever been to Earth before. Call warns Ripley for the military that will come looking for her soon. She says someone can get pretty lost on Earth, and asks Ripley what to do. Ripley replies that she doesn't know since she is a stranger there herself. A wide shot shows Call and Ripley sitting in the hills overlooking a desolate and ruined Paris.