Tom Eaves’ strike after 48 minutes put Town on their way, before Francois Zoko put the result beyond doubt on 89 minutes to see off the Young Royals and claim top spot in Southern Group A.
With two wins and a draw to their name, the Glovers secured top spot by two points ahead of Reading, whilst extending their unbeaten run in all competitions to seven matches.
Darren Way made four changes to the side that drew 2-2 with Solihull Moors in the Emirates FA Cup First Round at Huish Park on Saturday.
The suspended Kevin Dawson dropped out of the squad and was replaced by Matt Butcher who returned from injury in timely fashion, whilst Bevis Mugabi, Jonny Maddison and Tahvon Campbell replaced Darren Ward (rested), Artur Krysiak (on the bench) and Eaves (on the bench) respectively.
In an eerily empty Madejski Stadium, it was the Young Royals who produced the first chance of the game after five minutes when Andrija Novakovich cut inside from the left before curling an effort past Maddison’s far post.
Town responded with an effort of their own minutes later, when Matty Dolan stung the fingertips of Stuart Moore in the home goal with a driven shot from 25 yards.
That was as good as it got in the opening 25 minutes as Reading enjoyed plenty of possession with little by way of penetration, whilst Town looked to commit numbers on the counter-attack.
The Glovers tactic almost worked to full effect just before the hour mark, when a swift move led to Ryan Hedges cutting inside from the right and firing at goal, with the Welshman’s well-struck strike drifting wide of the target.
Reading continued to enjoy large spells of possession for the remainder of the half, but struggled to find a way past a resolute Glovers defence that had conceded just two goals in the last six matches coming into the fixture.
Danzell Gravenberch fired straight at Maddison from range, before Liam Kelly’s cross almost snuck in at the back post but, unable to produce a meaningful attempt on goal, both sides trudged in for half-time at deadlock.
The Glovers made a change at the break with Eaves replacing Campbell for Town, something which transpired to be an inspired substitution just three minutes after the restart.
From a deep free-kick, Mugabi won the initial aerial challenge with his flick-on landing at the feet of Eaves, who smashed home from close range with his first touch since taking to the pitch.
Buoyed by taking the lead Town enjoyed a good spell of possession at the Madejski, finding an easier outlet through the hold-up play of Eaves through the middle.
Despite seeing more of the ball it was the Young Royals who drew the game’s next chance, with substitute Josh Barrett firing into the arms of Maddison from inside the box after a slick move just past the hour mark.
That warning sparked an immediate response from the Glovers; Liam Shephard’s excellent run caused mayhem in the Reading box resulting in Ben Whitfield flashing a ball across the face of goal, before Eaves blasted an effort just wide from the edge of the box minutes later as Town looked to affirm their spot at the top of Southern Group A.
As the game entered the final 10 minutes Reading pressed for an equaliser which would see them regain top spot in the group, but Maddison was on hand to deny both Barrett and Sam Smith with smart saves.
The young stopper’s saves were to prove all the more important in the closing stages as substitute Francois Zoko put the game beyond doubt with a calm finish from six yards after Dolan’s free-kick ricocheted and fell to the Ivorian inside the box.
With just three minutes of added time to find a way back into the game Reading Under-23’s were once again denied by Maddison who produced an excellent low stop to deny the Young Royals.
The Glovers saw out the final few minutes to claim a deserved 2-0 win and qualify for the knockout stages top of the group, with an unbeaten record in the group stages.
Reading Under-23’s: Moore; Watson, Gravenberch, Cooper, McIntyre; Rinomhota, Kelly (Barrett 45’), Richards (Smith 79’); Frost, Samuel (Keown 63’), Novakovich.
Subs not used: Tupper, Andresson, East, Van Cooten.
Yeovil Town: Maddison; Shephard, Lacey, Smith, Mugabi; Whitfield, Butcher (Ezewele 81’), Dolan; Hedges, Campbell (Eaves 45’), Khan (Zoko 70’).
Bookings: Mugabi 42’
Subs not used: Krysiak, Copp, Sowunmi.