Pompey will play Crystal Palace U21s in the group stages of the 2021/22 Papa John's Trophy.
The Premier League side's youngsters were drawn into Southern Group B, which already contained the Blues, AFC Wimbledon and EFL newcomers Sutton.
Pompey will take on the Eagles and one of the other clubs at Fratton Park, while also having one away fixture as they battle for a place in the knock-out stages.
Kenny Jackett guided the Blues to EFL Trophy glory with a memorable penalty shoot-out victory over Sunderland in 2019, but the Blues were defeated by Salford in the delayed 2020 showpiece.
| Southern Group A | Southern Group B | Southern Group C | Southern Group D |
| Colchester | AFC Wimbledon | Burton | Forest Green |
| Gillingham | POMPEY | MK Dons | Northampton |
| Ipswich | Sutton | Wycombe | Walsall |
| West Ham U21 | Crystal Palace U21 | Aston Villa U21 | Brighton U21 |
| Southern Group E | Southern Group F | Southern Group G | Southern Group H |
| Bristol Rovers | Newport | Charlton | Cambridge |
| Cheltenham | Plymouth | Crawley | Oxford |
| Exeter | Swindon | Leyton Orient | Stevenage |
| Chelsea U21 | Arsenal U21 | Southampton U21 | Tottenham U21 |
| Northern Group A | Northern Group B | Northern Group C | Northern Group D |
| Carlisle | Oldham | Crewe | Bolton |
| Hartlepool | Salford | Shrewsbury | Port Vale |
| Morecambe | Tranmere | Wigan | Rochdale |
| Everton U21 | Leeds U21 | Wolves U21 | Liverpool U21 |
| Northern Group E | Northern Group F | Northern Group G | Northern Group H |
| Doncaster | Bradford | Accrington | Harrogate |
| Rotherham | Lincoln | Barrow | Mansfield |
| Scunthorpe | Sunderland | Fleetwood | Sheff Wed |
| Man City U21 | Man United U21 | Leicester U21 | Newcastle U21 |