Massimiliano Allegri's side suffered a surprise 2-1 defeat in the home tie last month, as Mohamed Salah scored either side of Fernando Llorente's strike.
Marchisio suffered a knee injury while on international duty and with Juve needing victory by two goals to progress to the final, Allegri hopes the midfielder will be able to play some part, with Andrea Pirlo (calf) and Paul Pogba (hamstring) still out.
"Marchisio is doing well," Allegri said on Monday. "He's returned to the squad, however he's been working separately.
"Considering the number of games he's played from the start of the season to now, I think four or five days out of the team did him some good.
"He'll be available, so I'll have to assess today whether to play him or not.
"As for Pirlo, the day after tomorrow he should be part of the group, and then we'll slowly evaluate his physical condition.
"I'll assess him once he returns to the group, but the important thing is to have him available. I hope he'll be in good condition, because he's done well in training.
"There are still a lot of games between now and the end of the season, I think Andrea has the quality to play in big games, even when he's not in top condition."
Allegri says Fiorentina start the second leg as favourites to progress, and has pledged to play a back four with an extra attacker, while Marco Storari will start in goal instead of Gianluigi Buffon.
Juve go into the game on the back of five straight wins across all competitions since the first-leg defeat, while Fiorentina have lost just once in their past 19 outings.
"It's a big opportunity for us," said Fiorentina coach Vincenzo Montella. "To beat Juve after also knocking Roma out would mean we'd eliminated the top two teams in the league, and that would be a source of pride for us.
"We're very proud, but we haven't achieved anything yet. Up until now we haven't reached any of our goals.
"Tomorrow's game can give us the opportunity to achieve something positive. Our state of mind is that we can achieve more and more, the city recognises our commitment and is proud of us."
Fiorentina are bidding to reach their second straight Coppa final, after losing 3-1 to Napoli in last year's edition, while Juve were beaten by the same opponents in the showpiece in 2012, last lifting the trophy in 1995.
