Holders Peterborough United have been drawn at home to Northampton Town in the last 32 of the newly-renamed Vertu Trophy.
Peterborough, who beat Wycombe in last season’s final, progressed from the group stage of this year’s competition with a 100% record, winning all three games.
Wycombe, the League One leaders, also have a home tie against Ian Holloway’s Swindon Town, while twice winners Port Vale, who are top of League Two, will travel to boss Darren Moore’s former club Doncaster Rovers.
Aston Villa Under-21s will be away to 2002 and 2004 winners Blackpool, while Wrexham, who lifted the trophy in 2005, will be at home to Crewe Alexandra.
Birmingham City – winners twice in the 1990s and playing in the competition for the first time since then – will make a long trip to Devon to play Exeter City.
All 16 group winners earned a home draw in the first knockout round and each will host one of the runners-up, the tournament remaining regionalised until the quarter-final stage.
Each group included one Premier League academy side but Villa were the only one to progress by finishing second in their group behind Bolton – who won the trophy for the second time in 2023 and will face Huddersfield Town – with the other 15 academy teams all eliminated.
The competition, predominantly for League One and League Two clubs, was known as the Bristol Street Motors Trophy up until earlier this week.
The last-32 ties will be played during the week beginning 9 December.
Vertu Trophy last-32 draw
Northern section
Draw A
Morecambe v Lincoln
Wrexham v Crewe
Stockport v Bradford
Bolton v Huddersfield
Draw B
Rotherham v Tranmere
Blackpool v Aston Villa U21
Chesterfield v Wigan
Doncaster v Port Vale
Southern section
Draw A
Wycombe v Swindon
Peterborough v Northampton
Charlton v Leyton Orient
Walsall v Reading
Draw B
Cheltenham v Cambridge
Colchester v AFC Wimbledon
Burton v Stevenage
Exeter v Birmingham