A shocking DNA comparison between 15th century English monarch Richard III and a set of descendants found a break in the 500-year lineage, which could throw off centuries of claims to the British throne, including much of the Tudor dynasty. Richard III’s DNA matched those of the descendants of his sister, Anne of York, but did not match descendants of the male side of his family.

Regardless of the potential breaks in lineage, Queen Elizabeth II is still the queen of England, because she is of the much later House of Windsor and her claim to the throne is in only the most distant way related to Richard III. The DNA tests also found that Richard, who is commonly thought to have had brown hair and greyish eyes, could have actually had blond hair and blue eyes. There is a 96 percent chance of him having blue eyes and a 77 percent chance of him being blond at some point in his life, likely during his childhood, according to the Guardian.

Some parts of DNA are passed down through maternal or paternal lines almost untouched, like mitochondrial DNA, which is handed down through the female line. Genetic researchers use these to tell if a person is related to another.

The Daily Mail has put together a family tree to connect Richard the III and two of the present-day descendants of Anne of York. Researchers are unsure where the break occurred in the line, but their results mean that at some point a child in the line was fathered by someone other than one of the men in the family tree, according to news.com.au. That break would be most significant if it occurred between Edward III (who died in 1377) and his son, John of Gaunt.

“John of Gaunt was the father of Henry IV, so if John of Gaunt was not actually the child of Edward III, arguably Henry IV had no legitimate right to the throne, and therefore neither did Henry V, Henry VI, and, indirectly, the Tudors,” said Kevin Schurer, pro-vice chancellor of the University of Leicester in England.

Richard III’s bones were found under a parking garage in Leicester in 2012. DNA tests in 2013 confirmed that the remains were his. He was killed in 1485 at the Battle of Bosworth, near Leicester, the battle that gave the Tudor family the English throne. He was the last English king to die in battle, according to the Guardian.