The five-time major winner was the star of a marquee group that also featured world number one Rory McIlroy and Bryson DeChambeau, shaking off an early bogey to card eight birdies and build a 13-under par total of 127.
He was one stroke in front of overnight leader Mackenzie Hughes of Canada and morning pacesetter Will Gordon.
Mickelson finished in style, nearly holing his approach shot at 18 and tapping in for a closing birdie.
It marks the first time in a career includes 44 PGA Tour titles that Mickelson has opened a tournament with two rounds of 64 or better.
“I’ve hit some good wedges, but more important than that I’ve given myself fair chances because I’ve hit a lot of fairways,” Mickelson said. “I felt like I was playing aggressive because of that.
“I also made some good putts today,” added Mickelson, whose birdies included three in a row at 13, 14 and 15 where he drained putts of six feet, 32 feet and seven feet.
“I made some tough six-footers that really kept the round going,” he said. “The birdies at 13 and 15 were not easy, there was a lot of break.”
Hughes, who torched TPC River Highlands for a 10-under 60 on Thursday, battled to a 68 for 128.
One-under through nine holes after three birdies and two bogeys, he made just one birdie on the back nine.
Gordon, making his eighth tour start on a sponsor’s exemption, made the most of his chance with an eight-under-par 62.
The 23-year-old American had nine birdies against a lone bogey in his first tour start since the Puerto Rico Open as the tour took a three-month hiatus starting in March because of the COVID-19 pandemic.
“I hit a little bit better today and the putter has been working really nice,” said Gordon, whose best PGA Tour result is a share of 10th in last year’s RSM Classic. “Just trying to stay out of my own way and stay in the moment. I’ve been really free and letting them go in.”
Gordon started on the back nine and birdied six of his first eight holes, the first of them on a 45-foot putt at the 10th.
He rolled in a 25-footer at the eighth for his final birdie of the day.
McIlroy had four birdies and two bogeys in a two-under-par 68 and headlined a group on nine-under 131 that also included Americans Xander Schauffele, Brendan Steele and Brendon Todd and Australian Marc Leishman.
Steele earned his share of fourth place with a sparkling 62. Todd and Leishman both signed for 65s while Schauffele carded a 68.
– Morikawa misses cut –
American Collin Morikawa, who made his first 22 consecutive cuts as a professional, missed the mark, ending the second-longest such streak in the last 30 years three shy of matching Tiger Woods with 25.
Morikawa opened with a 72 and only managed a 71 on Friday.
“It was bound to happen at some point,” he said. “Now I guess we’re going to stop talking about it.”
The tournament is the third since the PGA Tour resumed play, and saw another player test positive for COVID-19 on Friday after two caddies and a player tested positive on the eve of the tournament.
Denny McCarthy tested positive after feeling especially sore and tired after the first round. He went into quarantine while one first-round playing partner, Bud Cauley, withdrew as a precaution and another playing partner, Matt Wallace of England, opted to carry on.