London-born Rice played three times for the Republic but qualified for England as they were all friendly appearances. England host the Czechs on 22 March and are away in Montenegro on 25 March.
Rice, who has Irish grandparents, played for the Republic from under-16 level and went on to make three senior friendly outings last year, including games against France and the United States before the 2018 World Cup.
He broke into the West Ham first team at the start of the season and, as he considered his international future, was omitted from Republic squads because playing competitive games would have stopped him switching allegiance.
Why no Wan-Bissaka or Angus Gunn?
Southampton goalkeeper Angus Gunn and Crystal Palace full-back Aaron Wan-Bissaka are both overlooked by Southgate.
The in-form pair are dual-qualified – Wan-Bissaka has played once for the Democratic Republic of Congo at under-20 level and Gunn is coveted by Scotland.
Southgate said: “I could pick players, give them a cap and stop them playing for someone else but I don’t think that’s right. Angus has got a good challenge to get into our under-21s. There’s some good competition for places.”
Goalkeepers: Jack Butland (Stoke City), Tom Heaton (Burnley), Jordan Pickford (Everton)
Defenders: Trent Alexander-Arnold (Liverpool), Ben Chilwell (Leicester City), Michael Keane (Everton), Harry Maguire (Leicester City), Danny Rose (Tottenham Hotspur), Luke Shaw (Manchester United), John Stones (Manchester City), James Tarkowski (Burnley), Kieran Trippier (Tottenham Hotspur), Kyle Walker (Manchester City)
Midfielders: Ross Barkley (Chelsea), Dele Alli (Tottenham Hotspur), Fabian Delph (Manchester City), Eric Dier (Tottenham Hotspur), Jordan Henderson (Liverpool), Ruben Loftus-Cheek (Chelsea), Declan Rice (West Ham)
Forwards: Harry Kane (Tottenham Hotspur), Marcus Rashford (Manchester United), Jadon Sancho (Borussia Dortmund), Raheem Sterling (Manchester City), Callum Wilson (Bournemouth)