Dufner, who lost last year’s PGA Championship in a playoff to rookie Keegan Bradley, birdied three of his first five holes en route to a 10-under total of 132 at the Copperhead Course at Innisbrook in Palm Harbor, Florida.
Ireland’s Padraig Harrington (73), who was the overnight leader after setting the course record with a 61 on Thursday, was two shots back after mixing three bogeys with a birdie.
The Dubliner was tied in second place with American William McGirt (68), while England’s Luke Donald, the world No. 2, is one of five players sitting three shots off the pace.
Donald can reclaim his spot at the top of the world rankings from Northern Ireland’s Rory McIlroy with a win this week.
Spain’s Sergio Garcia (68) is also well-placed for a weekend push sitting just four shots back of Dufner along with Australian John Senden (70) and Americans Jim Furyk (70) and Kenny Perry (70).
For Dufner, who finally broke through on the PGA Tour last year, his second consecutive round of 66 indicates he could be hitting peak form just in time for the first major of the year at Augusta National next month.
But the 34-year-old Clevelander, who has just two wins on the Nationwide Tour to his name, will be aiming on Sunday for his first victory on the PGA Tour.
“It’s definitely a goal, I definitely felt like last year after some close calls I was ready to make that jump and start winning some tournaments,” said Dufner, who has gone bogey-free in his opening two rounds.
“It could be a great stepping stone and give me some more confidence going forward.”
Three-times major winner Harrington, searching for his top form after 18 difficult months sent him plummeting to 90th in the world rankings, said he paid the price for a conservative approach after a wonderful opening round.
“I’m happy with my game but when you shot a good score, it’s hard to match the next day,” said Harrington.
“When you’re leading the tournament, it’s hard not to be a little bit cautious and I think at times I was on the greens. I struggled to get the ball up to the hole a lot today.”
Among those missing the cut, which was set at one-under 141, were Japan’s Ryo Ishikawa, Colombian Camilo Villegas and Masters champion South African Charl Schwartzel, who followed up a five-over 76 on Thursday with a two-over 73 on Friday.
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In Jupiter, Florida, Tiger Woods returned to practice Friday and declared himself ready to play next week.
Woods said in a story on his website that his left Achilles tendon felt good and that he would play in the two-day Tavistock Cup exhibition that starts Monday, and then the Arnold Palmer Invitational at Bay Hill.
Woods withdrew from the Cadillac Championship on Sunday after 11 holes with what he described as tightness in his Achilles tendon. He said he didn’t want to risk further damage. One day later, he said doctors told him it was only a mild strain.
It was the left Achilles that caused him to miss two majors last year.
Woods has won Bay Hill six times. It will be his final tournament before the Masters begins April 5.
