The Bluebirds had an opportunity to go top of the Championship with a win but in the end they were forced to settle for a point.
Bothroyd, who had cut a rather forlorn figure in his return to the Cardiff line-up, saw his long-range strike come back off the underside of the bar though referee Andy D'Urso adjudged that the ball had crossed the line.
The goal cancelled out Noel Hunt's strike early on and in an end-to-end match, a draw was perhaps a fair result.
Cardiff were looking to bounce back from their defeat to local rivals Swansea on Saturday and boss Dave Jones opted to make five changes to his team.
The most notable of which saw the Championship's top scorer Bothroyd return to the starting line-up following a suspension in favour of the rested Craig Bellamy.
Royals manager Brian McDermott made two changes from the side that was beaten by QPR at the weekend with Simon Church and Hunt replacing Jay Tabb and Hal Robson-Kanu.
And it was those two players who combined to put the hosts in front in the fifth minute.
Church rode the challenges of several Cardiff defenders down the right channel before whipping a cross to the far post which Hunt was able to poke home.
The Bluebirds' Seyi Olofinjana thought he had equalised when he drilled the ball past Adam Federici but the Nigerian international had already been flagged offside.
Church had an opportunity to double his side's lead as Shane Long pulled the ball back to his fellow striker but the Welshman's drilled effort from 12 yards was magnificently saved by Tom Heaton.
Church might have been made to rue that miss but Michael Chopra spurned a glorious chance to draw Cardiff level.
The former Newcastle man managed to get in between the Reading defenders but when he found himself one-on-one with Federici, he fired wide.
On the stroke of half-time, Cardiff launched a rapid counter-attack and when Chopra squared the ball across the six yard box, Olofinjana looked certain to score with the goal at his mercy but he was superbly tackled by Royals defender Andy Griffin.
After the interval, the lively Long had a chance to put Reading further in front but his 20-yard lob went narrowly over the bar.
At the other end, Cardiff's best chance of the match came when Chris Burke cut inside on the right and dribbled the ball into the area but he struck wide.
The Bluebirds, galvanised by this chance, went to enjoy a good spell of pressure and they were rewarded for their efforts in contentious circumstances 13 minutes from time.
Chopra picked out strike partner Bothroyd and his 35-yard strike went in off the underside of the bar.
Reading were adamant that the ball had not crossed the line, nevertheless the goal stood.
Burke's 20-yard strike nearly won it for Cardiff at the death but Federici produced a fine save to ensure the spoils were shared.