Michael Long, the defending champion, posted a 2-under 70. He shares second place with John Douma (65), Brian McCann (68), Tom Scherrer (68) and Johnson Wagner (70) at 7-under-par 137.
Thompson coasted out of the gate on the back nine with six straight pars at the Tournament Players Club of Virginia Beach. He moved into red figures for his round with an eagle at the par-5 16th.
The 25-year-old ran off another stretch of six pars in a row around the turn. Thompson then dropped in a birdie at the par-5 fifth to move to minus-7. The South Carolina alum birdied the par-5 eighth to grab the overnight lead.
'I could be leading this tournament by a lot right now, but that's the way it goes,' said Thompson. 'I had so many putts that were just off the edge. I had a stretch of eight holes where I had six putts lip out. If I had been able to use my string, I would have needed about three inches to make them all.'
Long is in line to set tour history this week. No player ever on the Nationwide Tour has successfully defended his title.
Long parred the first six holes of his round before birdieing the 16th. He birdied the third to move to minus-7. Long dropped a shot at the seventh, but came back to birdie the eighth for a share of second place.
'I don't know about this defending thing,' said Long. 'I'm just trying to enjoy it. It's something where you try to do the best you can.'
Wagner, a Virginia Tech graduate, carded back-to-back birdies from the second and again from the seventh to climb to 9 under. However, he bogeyed the 10th and 13th before dropping in a birdie at the 15th. He slipped to a bogey at 16 before parring his last two holes.
Scherrer birdied two of his first nine holes and climbed to 7 under with birdies at the first and sixth. After a bogey at the seventh, the two-time Nationwide Tour winner birdied the eighth to share second place.
McCann carded one lone birdie over his first 13 holes. He then birdied the par-5 fifth and came right back to birdie the sixth. He jumped to 9 under with an eagle at the eighth, but McCann bogeyed the ninth to slide back one shot.
Douma dropped in birdies at the third and sixth, before an eagle at the eighth moved him to minus-4. He birdied the 11th, then ran off three consecutive birdies from the par-4 15th to jump to 9 under. He bogeyed the last to slip into a share of second. The bogey also cost Douma a share of the course record of 64, which four players have shot.
'Today came easy compared to yesterday,' Douma said. 'It was a grind to shoot par. Today, I got the putter working. I didn't make any long bombs. I hit some irons close and made some short ones, too.'
A total of 70 players made the cut, which fell at even-par 144. Among those who missed out on the weekend were Ryan Armour (145), Ben Bates (146), Keoke Cotner (146) and Robin Byrd (147).