The 46-year-old leaves the South Yorkshire outfit after nearly three seasons in charge and a 15th-place finish in the table last season.
Ferguson announced his decision in a statement released by the League Managers Association.
The statement read: "Following recent discussions and meetings with the board I have decided to resign from my position as manager of Doncaster Rovers Football Club.
"I'm disappointed to have made this decision as I have enjoyed my time at the club and was looking forward to the new season, but I felt I had no alternative.
"I would like to thank the board for the opportunity and I am grateful to the players, staff and fans for their support. I wish the club all the best in the future."
In a separate statement, Doncaster wrote: "Following conversations between the board of Doncaster Rovers and Darren Ferguson, the club can confirm that we have parted ways.
"The board intends to start the recruitment process for the new football manager immediately, and will provide a further update in 24 hours."
Ferguson secured automatic promotion from League Two with Doncaster in May 2017.