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| 1st(1st leg) | Sokol Živanice | - | FK Baumit Jablonec | 3:6 | ||
| 2nd (2nd leg) | FK Baumit Jablonec | - | Sokol Živanice | 5:1 | ||
| 1st(1st leg): 07/10/2009 15:00 | 2nd (2nd leg): 28/10/2009 13:30 | Winner: FK Baumit Jablonec | ||||
| 1st(1st leg) | 1. FK Příbram | - | Baník Ostrava | 2:0 | ||
| 2nd (2nd leg) | Baník Ostrava | - | 1. FK Příbram | 0:0 | ||
| 1st(1st leg): 07/10/2009 15:30 | 2nd (2nd leg): 27/10/2009 15:30 | Winner: 1. FK Příbram | ||||
| 1st(1st leg) | FC Hradec Králové | - | FK Teplice | 1:2 | ||
| 2nd (2nd leg) | FK Teplice | - | FC Hradec Králové | 2:1 | ||
| 1st(1st leg): 07/10/2009 16:00 | 2nd (2nd leg): 11/11/2009 16:00 | Winner: FK Teplice | ||||
| 1st(1st leg) | Bohemians 1905 | - | Slovan Liberec | 1:2 | ||
| 2nd (2nd leg) | Slovan Liberec | - | Bohemians 1905 | 1:1 | ||
| 1st(1st leg): 07/10/2009 17:00 | 2nd (2nd leg): 28/10/2009 15:00 | Winner: Slovan Liberec | ||||
| 1st(1st leg) | SK Kladno | - | Viktoria Plzeň | 0:3 | ||
| 2nd (2nd leg) | Viktoria Plzeň | - | SK Kladno | 1:1 | ||
| 1st(1st leg): 07/10/2009 17:00 | 2nd (2nd leg): 13/10/2009 17:00 | Winner: Viktoria Plzeň | ||||
| 1st(1st leg) | 1. SC Znojmo | - | Sigma Olomouc | 1:3 | ||
| 2nd (2nd leg) | Sigma Olomouc | - | 1. SC Znojmo | 0:0 | ||
| 1st(1st leg): 08/10/2009 15:00 | 2nd (2nd leg): 28/10/2009 15:30 | Winner: Sigma Olomouc | ||||
| 1st(1st leg) | Dynamo České Budějovice | - | AC Sparta Praha | 0:5 | ||
| 2nd (2nd leg) | AC Sparta Praha | - | Dynamo České Budějovice | 1:0 | ||
| 1st(1st leg): 13/10/2009 17:00 | 2nd (2nd leg): 12/11/2009 17:00 | Winner: AC Sparta Praha | ||||
| 1st(1st leg) | MFK Karviná | - | Slavia Praha | 1:0 | ||
| 2nd (2nd leg) | Slavia Praha | - | MFK Karviná | 2:0 | ||
| 1st(1st leg): 14/11/2009 13:00 | 2nd (2nd leg): 25/11/2009 16:30 | Winner: Slavia Praha | ||||
