It runs in the San Joaquin Valley from Interstate 5 south of Los Banos to State Route 99 in Turlock.

State Route 165 begins at the junction of Interstate 5 south of Los Banos and heads north on a rural two-lane highway known as Mercey Springs Road.

The highway crosses the California Aqueduct north of Interstate 5 and enters Los Banos several miles later.

After leaving Los Banos, the highway then runs right through the San Luis National Wildlife Refuge and skirts the Great Valley Grasslands State Park to the west while crossing the San Joaquin River.

[5] Except where prefixed with a letter, postmiles were measured on the road as it was in 1964, based on the alignment that existed at the time, and do not necessarily reflect current mileage.