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Twilight on the Rio Grande finds Gene Autry (Gene Autry) on vacation in Mexico with Pokie (Sterling Holloway) when they learn their buddy Dusty Morgan (Bob Steele) has been killed with a knife in the back. Gene rushes to investigate the murder with a little help from hot-tempered Elena Del Rio (Adele Mara). They find the murder is connected to a gang of jewel thieves and the trail takes them back across the Rio Grande to a border town. There, Gene, Pokie, Champ (Champion Jr.) and Elena try to round up the murderous gang. Interspersed with the fight scenes is Gene delivering several cowboy songs with his trio the Cass County Boys, including "Twilight on the Rio Grande," "The Pretty Knife Grinder," "It's My Lazy Day," and "The Old Lamplighter". The 1947 Republic film was directed by Frank McDonald. It was filmed on location in Joshua Tree National Monument and Victorville, California.