The defending champion had to sink a 15-foot par putt on the final hole to preserve his slender advantage and card a two-over 74 for four-under 140.
Earlier starter McIlroy also made a key par-saving putt on the testing 18th for a one-under 71, carrying crucial momentum as he bids to complete the career grand slam with victory at the 80th Masters.
"I'm very excited. This is my best position going into the weekend at Augusta for a long time. I hope I can battle away tomorrow," the Northern Irishman told Sky Sports.
He will play the third round Saturday with arch-rival Spieth, who charged to eight-under early in his round before giving up four shots to par in testing conditions as the swirling wind dried out the treacherous putting surfaces.
"I thought it was a couple of a strokes higher than I was looking for," Spieth told Sky Sports.
"I've obviously got Rory right on my heels so it's no easy task ahead," he admitted.
U.S. amateur Bryson DeChambeau looked set to be alongside McIlroy as he stood three-under playing the last in the same group as Spieth, heading also for the only round in the 60's on the second day.
But a horrid hook off the tee and some hashed attempts at getting out of trouble led to a triple bogey to slip back to level par.
New Zealand's Danny Lee and American Scott Piercy are tied for third on two-under after rounds of 74 and 72 respectively, with Brandt Snedeker, Dane Soren Kjeldsen and Hideki Matsuyama of Japan the only other players under par after a testing second round where the scoring average was over 75.
Big-hitting American Dustin Johnson has also moved into contention after matching McIlroy's 71 to join the seven-strong group at level par, four back off the lead, while world number one Jason Day is a further shot back on a crowded leader board.
With the 10-shot rule in place, 2012 and 2014 champion Bubba Watson narrowly survived on six-under, but Phil Mickelson and reigning British Open champion Zach Johnson will not be around for the weekend.
There was a sentimental farewell for 66-year-old Tom Watson, playing his final Masters, who battled hard to make the cut, but missed out by two shots.
After finishing with a par on the 18th, two-time champion Watson was given a rousing ovation by the Augusta patrons.
As expected, Ernie Els, who opened with a disastrous nine at the opening hole Thursday, also missed out.
He three-putted the same green in his second round for a double bogey, but showed his quality to card a 73, but nine-over overall.
The 46-year-old South African vowed to sort out his very public putting problems, but admitted it had been a difficult return to the course on the second day.
"It was a very weird, surreal feeling this morning. It was like I walked on to the range and even the players and caddies, they kind of just looked at me as if I don't have pants on or something. Not a nice feeling," he told gathered reporters.