It happened on Interstate 84 in Montgomery around 3:30 a.m., closing the eastbound lanes through Orange County for miles.
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The driver of the overturned big rig sustained a minor injury, while none of the construction workers were hurt.
The tractor-trailer overturned at Exit 5 on the interstate, which is being renumbered and will be the future Exit 28.
The rig hit numerous construction vehicles being used for the sign work.
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The incident closed eastbound I-84 for 10 miles, from Route 17 in Wallkill to Route 208 in Montgomery.
I-84 connections to Route 17 in Wallkill were also closed.
New York State Police are investigating the crash.
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