Mutant Ninja Turtles Gaiden takes place in an alternate universe branching off Eastman and Laird's Teenage Mutant Ninja Turtles Volume 1, and Splinter's backstory is the same as his Mirage counterpart up to sometime after Return to New York, Book Three but before the City at War arc, at a time when his sons were 17 years old.
In the Prologue, Splinter receives a postal package addressed to his long-dead master Hamato Yoshi. The package contains an old katana wrapped in a shroud. Splinter reads the enclosed scroll and leaves the package's contents in his room. He greets his sons Leonardo and Raphael, who have just returned home from the surface, and asks Raphael about his "stroll." But Raph had lied to his father and picked a battle with street thugs, and Leo caught him. Raphael tries to continue his lie, but Leonardo tells Splinter the truth, infuriating Raph. Splinter punishes Raphael by forbidding him from surfacing for the rest of the month. Shortly after, Splinter starts instructing Leonardo in a new spiritual technique, but is interrupted by a Shell Cell call from Donatello, who asks Splinter for the favor of acquiring and bringing home some spare parts. When Splinter returns home with Donatello's spare parts, he is horrified to find that his sons have retrieved the sword from his room and are trying to kill each other with it. Recognizing the sword as cursed, Splinter unsuccessfully tries to get Leonardo to drop the sword, but then realizes this is no longer possible. Recalling the package's enclosed scroll, Splinter explains that the curse can be neutralized by forcing the curse into someone's body and then killing them an object corrupted by it. He forces the curse into his own body, and commands Leonardo to use the cursed sword to kill him, which his son very reluctantly does. Splinter catches fire, and both he and the sword are destroyed and reduced to a pile of ashes.
Splinter's death was very traumatizing to his sons, and did not love to see them spend the next 17 years mostly failing to deal with their severe psychological scars. Splinter was not alive during this alternate universe's version of the City at War incident, nor did he see his surrogate daughter April O'Neil marry Casey Jones, and he never met his granddaughter Shadow Jones or Raphael's new mutant turtle student Renoir.
In Chapter 18: A Little Deeper in Donatello's World and Chapter 19: Fragments of Truth Inside His Mind, the reader learns that Splinter was later cloned by his son Donatello in his secret cloning lab none of his family knows about. This clone was flawed and did not survive, but was nevertheless the new vessel for the demon Yamata-no-Orochi. The Orochi's psychic presence deluded Donatello into believing the clone had lived, and is actually his father Splinter recuperating inside a giant jar. In reality, Donatello's ever-advancing cloning techniques are actually being manipulated by the demon to provide it a viable new host. To this end, Donatello clones himself and murders the clones again and again (delusionally believing the clones are his hated brother Leo) in an attempt to improve the quality of his cloning technology.
As an alternative universe version of the Mirage continuity's Splinter, this Hamato Splinter did not have substantially different relationships. But his final lessons to his sons are directly relevant to the difficulties they faced after his death, and continue to face 17 years later.
Donatello invented a virtual world simulator and proudly showed it to his father. But Splinter was concerned with his son's distaste for a difficult reality, and counseled him never to deny reality no matter how difficult it is.
Splinter counseled Leonardo that there may be a time when the family has to go their separate ways, and that it might be for the better. But Leonardo promised his father that if that happens, he would be the last one to leave.
Splinter praised Michelangelo of his ability to be happy in any situation, but cautioned him that someday that happiness could be shattered and he would have to pick up the pieces.
At a time when Raphael came to Splinter with existential questions and pessimism about their future, Splinter taught him that, as long as there is life, there is hope. In Splinter's final words, he told his sons that they are never alone, because they have each other.
- According to Tigerfog, Splinter's naming order is Hamato Splinter, to match the naming order used in the Mirage TMNT comics and 2003 TV series. However, all of Hamato Splinter's sons—Leonardo Hamato, Donatello Hamato, Raphael Hamato and Michelangelo Hamato—place the surname after their name in Western naming order. Nevertheless, Splinter also can be (and has been) called Splinter Hamato, including in certain MNT Gaiden website texts. (Theoretically, the same can be said of Yoshi Hamato and Saki Oroku, since most Japanese names have their naming order reversed in the English language anyway.)
- This incarnation of Splinter is slightly different from his Mirage incarnation in that he took the surname of his master Hamato Yoshi, and also passed it to his own four sons.
- Splinter being killed by one of his own sons is a nod to an unfinished Mirage volume 2 storyline, where Splinter receives a premonition foreseeing that one of his sons would ultimately kill him.
- Splinter being secretly cloned by Donatello was a plot point later reused in a canonical Mirage volume 4 storyline. But in that scenario, Donatello is not insane and is not trying to revive Splinter, but instead cloned the Splinter who had recently died at the Jones Farm, revealing that that Splinter had actually been a clone of the real Splinter.
- A comical variation of Splinter appears in Bakabakashii; see Picasso Splinter.