Manchester United 2 Reading 1
Man Utd: Nani 69', Hernandez 72'
Reading: McAnuff 81'
The defender was injured in a first-half tackle with Reading goalscorer Jobi McAnuff, and is a doubt for Saturday's trip to QPR.
"We'll have to assess it in the morning," said Sir Alex Ferguson, the Manchester United manager, after the match.
"It's an ankle injury obviously so I'd imagine he'll not be ready for Saturday anyway.
"But we'll see what he's like after that."
Reading goalkeeper Adam Federici, who made his name in the same fixture six years ago, was busy from the start at Old Trafford, after Antonio Valencia capitalised on a goalmouth scramble to head straight at the Australian.
The loss of Jones saw Nani being introduced, and his presence increased United's goal threat more, though it took until the 69th minute for the Premier League leaders to finally make their breakthrough.
Antonio Valencia provided a neat ball for the Portuguese, who drove home into the corner of the net to open the scoring.
Reading looked for an instant equaliser, with Adam Le Fondre only denied by good work from David De Gea, before Nani turned provider, crossing into the path of Javi Hernandez who evaded Sean Morrison to head home the second.
The Premier League relegation battlers refused to give up, however, and with nine minutes remaining McAnuff squeezed through the United back line to reduce the deficit.
And in a frantic final few minutes, both Le Fondre and Hernandez were denied.
"The last 10 minutes were a bit scary," said Ferguson.
"Some of the football tonight was fantastic and we could have scored a lot of goals. In that way I'm satisfied.
"We didn't want a replay though and in fairness, we dug in and defended the long throw-ins and the balls into the box.
"You cannot get any better than a home tie in the cup. It doesn't matter who we play, at Old Trafford we have got a chance."
The result now hands United a visit from either Middlesbrough or Chelsea in the next round.
"We gave them a real run for their money," he said.
"In the last 20 mins we played some very good football.
"But they have some fantastic players and Nani is one of them.
"It is the first time I have managed here and it was good for me personally and my players to come here.
"We have to come back here in the league and will know a bit more about what to expect.
"We really wanted to put on a performance for our fans and we have done that."