Goals from Jack Muldoon and Alex Pattison turned the Round of 32 Papa John's Trophy tie on its head as Town progressed to the next round of the competition with a 2-1 victory.
Having gone behind after just 11 minutes following Nicky Maynard's first time finish, Town responded well and in the second half dominated the contest with Jack Muldoon equalising nine minutes after the restart before Pattison got the winner 20 minutes before the end of play.
Town made just one change for the trip to Prenton Park with Ryan Fallowfield replacing the suspended Connor Hall, which saw Nathan Sheron deputise at centre half. Manager Simon Weaver also named just three players on the Town bench as Simon Power joined those already in the treatment room.
The opening chances of the Papa John's Trophy tie came for the home side when eight minutes into the contest, Mark Oxley was needed to keep hold of Callum McManaman's curling but tame effort after the ball fell for the former Wigan Athletic midfielder at the edge of the area.
Oxley was again called into action two minutes later when Nicky Maynard's deflected strike bounced off the keeper and away from goal. However, the follow up would lead to the opener as possession fell for Tranmere's right back Kyle Hayde, who placed a perfect cross to the feet of Maynard who found the back of the net with a first time finish.
The Town keeper was becoming a busy man in the opening stages as 10 minutes later he was then needed at full stretch to tip over Joe Maguire's header which looked destined to loop under the crossbar.
In recent weeks, Jack Diamond had looked the most likely threat for Town and it was proving to be the same against the Merseyside team. Keen to drive forward with the ball, the winger played a neat one-two with captain Josh Falkingham before taking aim at the Tranmere goal and narrowly missing the target.
The visitors were beginning to dominate possession as the game approached the half hour mark and when they didn't have the ball, were keen to win it back but nonetheless, Tranmere were still proving to be a threat and McManaman nearly doubled the host's lead when a long throw dropped for him no more than 10 yards out. However, he dragged his shot wide when he should have done better.
It would turn out to be the last real opportunity of the half and Weaver's side went into the dressing at the break with work to do if they wished to progress to the last 16.
The Town manager - who celebrated his 44th birthday on Monday - wasted no time in making a change as Jack Muldoon came into the fray at the start of second half, replacing Lloyd Kerry and seeing George Thomson move into centre midfield alongside Falkingham and Pattison.
Town's top scorer in the previous three seasons was straight into the action and just nine minutes after the restart, he got his side a deserved equaliser. Town had been passing the ball with pace and purpose at the start of the second half and it was Falkingham who chose to make the killer pass to the substitute Muldoon.
The striker had placed himself nicely on the right side of the penalty area to then swivel with the ball and prod his effort past Murphy's near post to level the score on the 54th minute.
Now in the ascendancy, Town looked to get in front for the first time in the contest and both Pattison and Lewis Page found the confidence to shoot from distance although neither could trouble the Tranmere keeper.
Just after the 70th minute Town did take the lead and it came thanks to Alex Pattison. The majority of hard graft had been done by Luke Armstrong who nicked possession just outside the Tranmere box and was through one on one with the Tranmere keeper. The striker looked to round the keeper, but Murphy read his movement only to parry the ball out to Pattison who was on hand to smash the ball into an empty net.
Tranmere reacted immediately with a triple substitution, but Town looked a side willing to throw every body on the line to keep their lead intact.
The home side had an effort from range through Calum MacDonald, but it was the visitors who nearly scored the fourth goal of the contest as Armstrong's header from a corner rattled off the cross bar.
In the 95th minute, Oxley - who had been a stand out player all game - pulled off a critical save to deny Elliot Nevitt who had reached a flick on. That would turn out to be the final bit of action as Town progressed to the last 16 of the Papa John's Trophy for the first time.
Town: Oxley, Fallowfield, Page, Falkingham, Burrell, Thomson, Sheron, Pattison, Kerry (Muldoon 46), Diamond, Armstrong
Subs not used: Cracknell, Orsi,
Goals: Muldoon 54', Pattison 71'
Cards: Thomson 76'
Tranmere: Murphy, Hayde, Davies, Maguire, MacDonald, Merrie (O'Connor 72), Watson, Duffy (Foley 62), Feeney (Jolley 72), McManaman, Maynard (Nevitt 72)
Subs not used: Doohan, Clarke, Walker.
Goals: Maynard 11'
Referee: Samuel Barrott