The Royals are still fifth in the table, but a poor run of form means that they are now just two points ahead of seventh, while the Millers are in trouble at the other end of the table in 22nd.
Reading lost 1-0 away to bottom of the table Wycombe Wanderers on Tuesday night, despite the Chairboys having 10 men for the final 25 minutes.
It was a fourth defeat in five games for Veljko Paunovic's side, who have gone from having aspirations of getting into the top two a few weeks ago to now looking nervously over their shoulders at the teams chasing the playoffs.
The usually clinical Royals have not scored in the last two matches, but they have not gone three successive league games without registering since December 2018.
Including Saturday's trip to the New York Stadium, Reading's next six games are all against sides in the bottom half, but they should take nothing for granted after Tuesday night.
Their away form has been better than their record at home of late, with the loss at Adams Park being their first in six matches on the road.
Rotherham have won just one of their last six home games as their form has suffered a dip following a mini-revival at the start of 2021.
The Millers are currently on a four-game losing streak, but they have not suffered five consecutive league defeats since April 2017.
Their latest loss came on Tuesday night as Nottingham Forest beat them 1-0 at the New York Stadium, with Paul Warne admitting that his side "looked jaded".
Going into the weekend, Rotherham are two points adrift of safety and will have one eye on next week's crucial Yorkshire derby against Sheffield Wednesday, who are one point and one place below them.
The Millers have only been victorious in one of their last nine meetings with Reading, with the Royals winning 3-0 back in October thanks to a brace from Yakou Meite and a penalty from Lucas Joao, which has proved to be Rotherham's heaviest defeat of the season to date.
Rotherham United Championship form: WWLLLL
Reading Championship form: DLLWLL
After conceding that fatigue has played a part in the Millers' recent poor run, Warne could make changes and restore players such as Kieran Sadlier, Shaun MacDonald and Freddie Ladapo to the starting XI.
Joe Mattock is back in training but will not be ready to face the Royals.
Reading boss Paunovic is hopeful that Yakou Meite might be available for the game, revealing that the forward is in "his final stage" of recovery after three weeks out with a groin problem.
Rotherham United possible starting lineup:
Johansson; Olosunde, A. MacDonald, Robertson, Harding; Wing, S. MacDonald, Sadlier; Ladapo, Smith, Crooks
Reading possible starting lineup:
Rafael; Yiadom, Morrison, Moore, Richards; Rinomhota, Laurent; Meite, Olise, Ejaria; Joao
We say: Rotherham United 0-2 Reading
Both teams will be looking at this fixture as the perfect time to play each other, and we could see a competitive game at the New York Stadium. If Paunovic is able to get a reaction from his players after Tuesday night, Reading should get the better of a tired Rotherham side.