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Kingsgate Native claimed victory by a length and three-quarters
Kingsgate Native became the only British-trained Group One winner at Royal Ascot this year after triumphing in the Golden Jubilee Stakes.
Favourite Takeover Target and Fat Boy made the early running but Fat Boy dropped away with two furlongs left.
Kingsgate Native and War Artist swallowed up the leader before Seb Sanders stretched clear on the John Best-trained 33-1 shot.
Sir Gerry got up to take third from the fast-tiring Takeover Target.
Maidstone trainer Best, who saddled shock 100-1 winner Flashmans Papers on the opening afternoon, said: "It means so much to the yard to have winners and to have winners here is the ultimate.
"Everyone has worked so hard at home and it's so good to do it on the day.
Kingsgate Native is going to be a star sprinter for a while
Winning jockey Seb Sanders
"I thought 33-1 was a crazy price for him. He won the Nunthorpe last year and shouldn't have been anywhere near that price.
"The problem we had was that last year he was a very mature two-year-old and I felt that the weight-for-age allowance was in his favour as a two-year-old. I wasn't sure whether that would translate as a three-year-old."
Jockey Sanders said: "He was a bit keen early and it just took me a while to get him into his rhythm and settle him.
John Mayne, Seb Sanders and John Best talk to Rishi Persad
"He carried me very easily to the two-pole and it was just a case of getting past those hardened horses in front.
"He knuckled down and picked up really well - he's given me a terrific ride throughout. He's going to be a star sprinter for a while."
Earlier the Queen celebrated her first win at Royal Ascot since 1999 when Free Agent landed the listed Chesham Stakes to hand the monarch her 20th Festival winner during her reign.
She was caught on camera shaking her hands in delight and then jokingly swooning as she left to greet the Richard Hannon-trained horse in the winner's enclosure.
"The Queen was very gracious, when I returned on the winner. This is as big as it gets - to ride a winner for the Queen is special," said winning jockey Richard Hughes.
Big Timer stemmed the tide for high numbers when winning the Wokingham Stakes from stall 28.
Linda Perratt's charge came with a strong run on the far side in the final furlong under Tom Eaves to deny Beaver Patrol, who was on the opposite side of the track in stall two, by half a length.
King's Apostle was a nose behind in third on the far side with Knot In Wood fourth.
Ireland's Aidan O'Brien wound up as champion trainer at the meeting with six winners in all after 11-8 favourite Macarthur won the Hardwicke Stakes and 7-4 favourite Honolulu captured the closing Queen Alexandra Stakes.
Stable jockey Johnny Murtagh rode all six of the O'Brien horses during the five days, including Yeats in the Gold Cup and three other Group One winners, to secure the top rider title.
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