The novel begins with an unnamed narrator's arrival in Medicine Bow, Wyoming, from back East and his encounter with an impressively tall and handsome stranger. The stranger proves adept at roping horses, as well as facing down a gambler, Trampas, who calls him a sonofabitch. (At the time, the word was an unacceptable insult in any society, except between joking friends.) The stranger lays a pistol on the table and gently threatens "When you call me that, smile!" . Known only as the Virginian, the stranger turns out to be the narrator's escort to Judge Henry's ranch in Sunk Creek, Wyoming. As the two travel the 263 miles to the ranch, the narrator, nicknamed the "tenderfoot" and the Virginian begin to come to know one another as the Tenderfoot slowly begins to understand the nature of life in the West, which is very different from what he expected. This meeting is the beginning of a lifelong friendship and the starting point of the narrator's recounting of key episodes in the life of the Virginian.
The novel revolves around the Virginian and the life he lives. As well as describing the Virginian's conflict with his enemy, Trampas, and his romance with the pretty schoolteacher, Molly Stark Wood, Wister weaves a tale of action, violence, hate, revenge, love, and friendship. In one scene, the Virginian is forced to participate in the hanging of an admitted cattle thief, who had been his close friend. The hanging is represented as a necessary response to the government's corruption and lack of action, but the Virginian feels it to be a horrible duty. He is especially stricken by the bravery with which the thief faces his fate, and the heavy burden that the act places on his heart forms the emotional core of the story.
A fatal shootout resolves the ongoing conflict with Trampas, after five years of hate. After Trampas shoots first in a duel, The Virginian shoots Trampas (in self defense) and leaves to marry his young bride. The Virginian and Molly ride off together to spend a month in the mountains and then journey back East to Vermont to meet her family. They were received a bit stiffly by the immediate Wood family, but warmly by Molly's great-aunt. The new couple return to Wyoming and the Virginian is made a partner of Judge Henry's ranch. The book ends noting that the Virginian became an important man in the territory with a happy family.
In 1908 Owen Wister defends the actual Virginian "Charles D Skirden" who at this point is a police officer in Philadelphia, PA. Charles Skirden is accused of murder of John Bradley, a member of the Shamokin Street Gang in November 1908. Bradley is the son of James Bradley and Letitia Gallagher (Daughter of Squire Patrick Gallagher) He is arrested at the family's behest, but Owen Wister and others come to Skirden's aid. Afterwards Charles Skirden, in 1911 takes charge of a game preserve in Long Island.