Goals from Suk Hyun-jun and Kwak Tae-hwi ensured that the Czechs suffered a 2-1 defeat.
The Czechs suffered a 2-1 defeat by South Korea in their final test before the European Championship, playing the last half hour with 10 men.
The Czechs are counting on a mixture of youth and experience to get them out of a tough Euro 2016 Group D in France that includes defending champions Spain as well as Croatia and Turkey.
Spain trounced the South Koreans 6-1 in their friendly on June 1.
The Czechs were upset 25 minutes into the first half when goalkeeper Petr Cech failed to clear Yoon Bit-garam's shot from a free kick into the top right corner.
Czech skipper Tomas Rosicky, fired just wide from a nearly identical position a few minutes later.
Playmaker Rosicky, 35, missed half of the Czechs' qualifying campaign and almost the whole season for Arsenal through injury.
And it was Rosicky who lost the ball in his own half, inspiring a quick break and a goal by striker Suk Hyun-jun, who blasted a shot from close range to give Korea a two-goal lead five minutes before the break.
The Czechs had several decent opportunities but they were snuffed out by the Koreans, who were able to get to the ball in one-on-one situations and threatened the goal from quick breaks forward.
The home side had a better start to the second half, Marek Suchy taking a shot from distance that was deflected into the net by Korean Kwak Tae-hwi just after the interval.
But the Czech revival was stemmed an hour into the game when defender Theodor Gebre Selassie hit an opposing player in a mid-air clash, earning a second booking and a sending off.
"More than the red card I was bothered by the first half...I was disappointed by that," coach Pavel Vrba said. "Certainly our substitutions helped, the game became much faster."
The Czechs play Spain in their first Euro 2016 match on June 13.
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