He added a 40-foot birdie putt at the penultimate hole en route to an eight-under-par 63 that matched his career-low round on the PGA Tour.
"I played what I would consider the scorable holes really well, and then I threw in some others," the 42-year-old told PGA Tour Radio.
"That's what happens on a day when you get in the low 60s. Everything kind of meshed together."
With an 11-under 131 halfway total, Dufner headed fellow Americans Max Homa (63) and Joel Dahmen (66) by one stroke with half the field back in the clubhouse.
McIlroy seemed likely to remain close to the lead until he butchered the easy par-four eighth, his 17th, where his tee shot found a bunker, from where he took five more strokes to negotiate the final 55 yards.
He also bogeyed his final hole and signed for a 70 that left the two-times Wells Fargo winner five shots off the pace.
Justin Rose was more pleased with his morning's work, a 67 to trail by six shots.
Dufner, unhappy with the state of his stagnant career, shook things up at the end of last season by changing his swing, putting stroke, equipment and caddie.
He has not had a top-60 finish all year, and has slipped to 230th in the world rankings, but Friday gave him hope things are finally turning in the right direction.
"I'm just settling in to playing some better golf," said the five-times PGA Tour winner.
(Reporting by Andrew Both in Cary, North Carolina; Editing by Christian Radnedge)
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