Rovers bowed out of the Liverpool Senior Cup on Tuesday night after a 1-0 defeat against Southport at the Merseyrail Community Stadium.
Southport started the brighter of the two teams as after just 30 seconds on the clock, Dion Charles broke into the Rovers box and fired over the bar.
Charles was again the danger man as he picked up a loose ball 30 yards from goal, ran at the Rovers back line but then dragged his shot a few yards wide of the mark.
On the 11th minute, Liam Edwards met an in swinging corner for the home side but his firm header failed to trouble Pilling.
Half way through the first half, Charles found space on the left and cut the ball back into a dangerous area inside the box but none of his onrushing team mates could latch onto it to get their shot off.
Shortly after, Rovers' first effort off the half came via Bailey Thompson. The striker cut inside on his right foot and rattled a shot off the crossbar from 20 yards.
It was all square heading into half time.
Tranmere came out of the traps quicker in the second half. Harvey Gilmour forced a corner which was played short however the following cross was cleared away from danger.
Dion Charles was Southport's biggest threat and that continued into the second half. He drove into the box, cut onto his right foot and forced a clever save by Pilling down to his right.
The next chance fell the way of Harvey Gilmour for Rovers on the 58th minute. Walker-Rice's dangerous free kick found the head of Gilmour but he diverted his header a few yards wide of the post from eight yards.
Getting into the closing stages of the game it was Charles again who looked most likely for the Sandgrounders. This time his left footed effort from the edge of the box whistled past the left hand post.
The deadlock was finally broken just three minutes from time. An unmarked Jason Gilchrist nodded the ball home from six yards out to give the home team the advantage.
Despite the best efforts of Tranmere, Southport kept the ball well in the final minutes and advanced through to the next round of the Liverpool Senior Cup
Tranmere team: Pilling, Kerr (Burton 88'), Bakayogo, Devine, Long (Nugent 55'), Gumbs, Walker-Rice, Spellman, Thompson, Gilmour, Salkeld. Unused subs: Passant, Hayde, Blackham