Shorts III left practice on Thursday and was limited on Friday, leading coach Gus Bradley to rule him out.
The 26-year-old, who has led the team in receiving yards the last two seasons, missed the team's first two preseason games with the hamstring injury but returned for 15 plays in the third game against Detroit. He said afterward he felt 100 percent and was ready for the regular season.
Shorts III had 66 catches for 777 yards and three touchdowns in 13 games last season, when he was hampered by a groin injury. He had 979 yards and seven TDs in 2012.
Shorts III is the most-experienced receiver on the team, but this will be his 12th missed game since the Jaguars drafted him in the fourth round in 2011.
- Chris Mascaro