"The warrant on Mr. Karr has been dropped by the district attorney," Karr's
lawyer Seth Temin told reporters.
"They are not proceeding with this case. We are deeply distressed by the fact that they took this man and dragged him here from Bangkok, Thailand with no forensic evidence confirming the allegations against him and no supporting factors leading to a presumption that he did anything wrong," Mr Temin said.
"The DNA associated with the victim in this case does not match John Mark Karr," District Attorney Mary Lacy said later.
"The family of Mr Karr cooperated by providing circumstantial evidence that Mr Karr spent Christmas with his family in Atlanta, Georgia," at the time of the 1996 murder, he said.
After initially say that they would release Karr, 41, the Boulder County sheriff has detained him to face child pornography charges.
The announcement that no charges would be brought against Karr came just hours before he was due for his first Colorado court appearance.
Legal experts said it would be difficult to bring the case to trial without a DNA match as scepticism grew over Karr's claims to have been with the former Little Miss Colorado when she died.
Evidence disclosed
Prosecutors have disclosed none of their evidence against Karr who, after his arrest in Thailand, professed his love for child beauty queen JonBenet and said she died by "accident".
Under Colorado law, formal charges usually must come within 72 hours of a defendant's initial court appearance.
The DNA found on the little girl's underwear was identified as belonging to a white male but has never been matched to a suspect in the murder, which for 10 years has baffled police and fascinated Americans.
JonBenet's parents, who were once said by authorities to be under an "umbrella of suspicion" in their daughter's death, were also excluded from having left the DNA. Her mother died of cancer in June this year.
Other evidence at the crime scene included a footprint found near JonBenet's body in the basement of her home and a palm print on a nearby door.
JonBenet's father, John Ramsey, found the girl's battered body in the basement on December 26, 1996, about seven hours after her mother stumbled on a bizarre letter claiming she had been kidnapped and demanding a US$118,000 ransom.
JonBenet had been strangled to death with a garrotte made from cord and a paint brush found in her mother's paint supplies, her skull fractured and her mouth duct-taped. Forensic evidence suggests she had been sexually assaulted.
Members of Karr's family insist he was out of state at the time of JonBenet's murder and could not have been involved in the crime.