Subaru Hasegawa has the ambition to become one of the best basketball superstars of all time. However, this comes to an apparent halt when his school's basketball team ceases activities for a year. Shocked by the news, he decides not to involve himself in basketball anymore.

One day, his aunt Mihoshi Takamura invites him to coach the Keishin Academy girls’ basketball team on how to play the sport, and he reluctantly agrees. He meets sixth-graders Tomoka Minato, Maho Misawa, Airi Kashii, Hinata Hakamada and Saki Nagatsuki, and begins training them in the sport with one goal: to defeat all strong opponents along the way.

Ro-Kyu-Bu! follows the effort of five elementary girls and their coach as they their cute style of playing basketball.