Grimsby Town's hopes of progressing in the Checkatrade Trophy remain in the balance after they were beaten by Doncaster Rovers in their second group game.
Goals in either half from Alfie May, the second of which came from the penalty spot, sent the Mariners to defeat at the Keepmoat Stadium.
Town boss Michael Jolley named a side showing seven changes from the XI that began the weekend's league victory over Port Vale, with Reece Hall-Johnson, Alex Whitmore, Charles Vernam and JJ Hooper the men to start both games.
Sam Russeell began in goal, with Hall-Johnson, Whitmore, Paul Dixon and Martyn Woolford in defence.
Jake Hessenthaler, John Welsh, Ben Pringle and Vernam were the midfielders, with Hooper and Ahkeem Rose in attack.
The first chance of the game went to the hosts, with Alfie Beestin blasting over the bar from close range on eight minutes, before a long-range shot from May was pushed away for a corner by Russell.
From the flag-kick, another effort from the home side was saved by Russell, with Rose in the right place to block the follow-up on the line.
The Town keeper was called into action again on 17 minutes to keep out a close-range strike from Jermaine Anderon, then an opportunity for the Mariners was not taken as Rose headed over the bar.
A good run down the right by Vernam on 21 minutes led to a low cross for Rose but the striker was off-target, shooting wide.
Then another chance for Town came after Vernam got in behind the Doncaster defence but saw his shot comfortably stopped by home keeper Ian Lawlor.
The breakthrough came for Doncaster on 23 minutes, with MAY letting fly with a shot from distance that beat Russell and flew into the top corner of the net.
Town's response came through a chance for Pringle just after the half-hour as he turned and shot after a cross from Hooper, although the ball went just wide.
Russell had to make a good save to stop May from getting his second of the game and with no further chances coming before the interval, the Grimsby deficit remained at one goal.
No changes were made before the restart and Town were first to attack in the second half, with a Hooper volley two minutes in flying wide of the target.
Pringle went closer to an equaliser two minutes later after a Hooper cross to the far post, although Lawlor got back to push the midfielder's shot away.
The visitors continued their good start to the half when they created another chance on 52 minutes but Hooper's drive from the edge of the box was stopped by the keeper.
But when the game's second goal came on 56 minutes, it was the hosts who doubled their advantage. MAY was brought down inside the penalty area by Dixon and got up to beat Russell from the spot to make it 2-0.
Will Longbottom came on for Beestin as Doncaster made the first substitution just after the hour, before a powerful left-footed shot from Vernam was saved.
Longbottom had a chance to make it 3-0 but his first-time shot went across the face of goal and wide of the far post.
The first Town change saw Louis Robles replace Vernam on 68 minutes, before Mitch Rose came on for Ahkeem Rose nine minutes later.
More substitutions were made by Doncaster, with Cody Prior replacing Shane Blaney, before James Morris came on for May with six minutes to go.
And the final Town change saw Brandon Buckley come on for his debut in place of Hooper with four minutes left on the clock.
Pringle was shown the yellow card after a trip on Herbie Kane, before Doncaster's Tom Anderson followed him into the referee's notebook for a foul on Robles.
Three added minutes were played at the end of the game, with the home side seeing the remainder of the match out to ensure they drew level with the Mariners on three points from their two group fixtures so far.
Both remain behind Newcastle United U21, who top the group and will visit Blundell Park in the final round of fixtures next month before the knockout stages.
Town now have a weekend without a league game following the postponement of their trip to Mansfield, meaning they are next in action at home to Exeter City on October 20.
1 Ian Lawlor
28 Mitchell Lund
35 Branden Horton
12 Tom Anderson (C)
25 Shane Blaney
22 Jermaine Anderson
15 Herbie Kane F
11 Ali Crawford
19 Alfie May
16 Paul Taylor
14 Alfie Beestin
13 Marko Marosi
2 Niall Mason
38 Anthony Greaves
32 Cody Prior
39 Rieves Boocock
21 Will Longbottom
34 James Morris
23 Sam Russell
2 Reece Hall-Johnson
3 Paul Dixon
7 Jake Hessenthaler
25 Alex Whitmore
30 Ben Pringle
16 Martyn Woolford
4 John Welsh (C) F
18 Charles Vernam
9 JJ Hooper
21 Ahkeem Rose
28 Ollie Battersby
26 Brandon Buckley
15 Harry Clifton
31 Matthew Pollock
8 Mitch Rose
29 Rumarn Burrell
14 Louis Robles
Attendance: 1,420 (143)
Referee: Michael Salisbury