Juventus star Marchisio was stretchered off 15 minutes into the champions' 4-0 victory over Palermo in Turin after a challenge that left him rolling on the turf in pain.
Juve's victory took their Serie A lead over Napoli to nine points with five games remaining, but the shine was taken off their achievements by the setback to Marchisio.
"We suspect it is his cruciate (ligament) but we'll wait for the results tonight or tomorrow," Juventus coach Massimiliano Allegri told Mediaset.
"We hope it isn't too serious."
If confirmed, Marchisio will be ruled out for the rest of the season and would, in all likelihood, miss this summer's football showpiece in France.
Juventus are aiming for a fifth consecutive league title, a feat they last achieved in 1935.