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Teenager Jordan Ball scored on his senior debut to give Doncaster a 1-0 win and knock JPT holders Chesterfield out of the competition.
Ball's first act after being introduced as a substitute after 83 minutes was to head home David Cotterill's deep corner and book his side's place in the next round.
It had been a game of few clear-cut chances with neither appearing in any rush to win the tie.
Chesterfield appeared happy to sit back and let Doncaster dominate possession for long periods of the game and proved very difficult to break down.
Visiting keeper Richard O'Donnell was forced into a number of saves without being greatly tested, with Robbie Blake turning a first-half header over from two yards in the best of the first-half chances.
The game opened up somewhat after the break but Doncaster continued to craft the better opportunities with Cotterill and Martin Woods trying their luck from distance before Ball won it.