The 2016 UniCredit Czech Open is a professional tennis tournament played on clay courts. It is the 23rd edition of the tournament which is part of the 2016 ATP Challenger Tour. It took place in Prostějov, Czech Republic between 31 May and 4 June 2016.
| Country | Player | Rank1 | Seed | |
|---|---|---|---|---|
| Ričardas Berankis | 50 | 1 | ||
 Czech Republic | Lukáš Rosol | 59 | 2 | |
| Czech Republic | Jiří Veselý | 60 | 3 | |
 Spain | Íñigo Cervantes | 69 | 4 | |
 Bosnia and Herzegovina | Damir Džumhur | 73 | 5 | |
 Kazakhstan | Mikhail Kukushkin | 83 | 6 | |
 Brazil | Rogério Dutra Silva | 85 | 7 | |
| Robin Haase | 86 | 8 |
The following players received wildcards into the singles main draw:
Dominik Kellovský Zdeněk Kolář
Alex MolčanThe following players received entry as alternates into the singles main draw:
The following players received special entry into the singles main draw:
The following players received entry from the qualifying draw:
The following players received entry as lucky losers into the singles main draw:
| Country | Player | Country | Player | Rank1 | Seed | |
|---|---|---|---|---|---|---|
| Nicholas Monroe |  New Zealand | Artem Sitak | 119 | 1 | ||
| Brazil | Marcelo Demoliner |  Austria | Julian Knowle | 131 | 2 | |
| Julio Peralta | Hans Podlipnik | 131 | 3 | |||
 Belarus | Aliaksandr Bury | Slovakia | Igor Zelenay | 138 | 4 |
The following pairs received wildcards into the doubles main draw:
Dominik Kellovský / Jaroslav Pospíšil Zdeněk Kolář / Matěj Vocel Michal Konečný / Václav ŠafránekSee main article: 2016 UniCredit Czech Open – Singles.
Mikhail Kukushkin def. 
Márton Fucsovics, 6–1, 6–2See main article: 2016 UniCredit Czech Open – Doubles.
Aliaksandr Bury / Igor Zelenay def. Julio Peralta / Hans Podlipnik, 6–4, 6–4
