Pliskova failed to convert two set points in the second set and lost 6-1, 7-6 (2).
It was the first time since 2008 that the top-seeded woman lost in the second round at the Open.
Pliskova was a finalist at the Open in 2016 for the best Grand Slam result of her career.
Garcia, ranked 50th, beat Pliskova for the fourth time in their seven meetings.
Li beat No. 13-seeded Alison Riske in a matchup between Pennsylvanians, 6-0, 6-3. Li, who is 20 and ranked 128th, now has two career victories over players ranked in the top 75, and the wins have come in her two matches this week.
Rogers beat No. 11-seeded Elena Rybakina 7-5, 6-1. By advancing to the third round, Rogers matched her best showing at the U.S. Open. She’s ranked 93rd.
Earlier in the day, Angelique Kerber moved into the third round with a solid showing that brought some much-needed confidence. Her 6-3, 7-6 (6) victory over Anna-Lena Friedsam had her feeling more at home in the place where she made her WTA Tour breakthrough in 2011 by reaching the semifinals.
“It’s a really special place for me,” the 32-year-old Kerber said. “Everything starts for me here in 2011, a long time ago.”
She hadn’t played competitively in seven months, which weighed on her when she headed for New York. How would all that time away from competition translate to a Grand Slam? How would it play out amid all the changes brought about by the coronavirus pandemic?
“I was sitting on the plane and I was like, ‘OK, let’s see what happens,” she said. “It’s not so easy. We had such a long break. You need a little time to find your way and your rhythm again.”
Kerber’s win with the roof closed at Louis Armstrong Stadium started the third day of the tournament, when a chance of rain was forecast.
No. 6 seed Petra Kvitova also moved into the third round with an uneven match that helped her adapt to the new conditions. She was nervous in the first set but played much steadier in a 7-6 (3), 6-2 victory over Kateryna Kozlova, ranked No. 99 in the world.
“With my nerves, I wasn’t moving well,” she said. “That’s a Grand Slam. At the beginning of the tournament it’s always like that.”
The US Open has been a big challenge for Kvitova, making her 14th appearance. It’s the only Grand Slam event where she has failed to reach the semifinals. She won Wimbledon in 2011 and 2014, but her best showings at Flushing Meadows were in 2015 and 2017 when she reached the quarterfinals.
The women’s bracket had one puzzling result.
No. 30 seed Kristina Mladenovic served for the victory while leading 6-1, 5-1 against Varvara Gracheva — and lost. She failed to convert four match points and let it slip away completely, with the No. 102-ranked Gracheva rallying to a 1-6, 7-6 (2), 6-0 victory in her first Grand Slam tournament.
Said Mladenovic: “She was brave and she went for it.”
The Associated Press contributed to this report.