Wadham was the first woman who was not a member of the royal family or titled aristocracy to found a college at Oxford or Cambridge.

Dorothy was the second and eldest surviving child of the very wealthy Sir William Petre (c.1505–1572), Secretary of State to four successive Tudor monarchs (namely Kings Henry VIII, Edward VI and Queens Mary I and Elizabeth I), who had acquired much property following the Dissolution of the Monasteries.

Part of the Petre inheritance received by Dorothy came from grants made by Queen Mary to her father Sir William Petre, of lands formerly held by Lady Jane Grey and forfeited to the crown, which had come in part from the great heiress Cecily Bonville, of Shute, Devon.

Later in life, her writing skill and knowledge of Latin was evident, and it is likely that she was educated at her home, Ingatestone Hall, Essex.On 3 September 1555 at St Botolph, Aldersgate, in the City of London, she married Nicholas Wadham (1531–1609).

His wishes included the founding of a college in Oxford University, and this Dorothy accomplished, noting that "it would greatly offend my conscience to violate any jot of my husband's will".

Dorothy managed to loosen Davis's ties by way of a collusive suit in chancery in July 1610, which established a trust excluding him.

The appointment of the Warden, Fellows, and Scholars, and even on occasion the college cook, rested with Dorothy, as shown by a series of letters written by her business agent John Arnold, and signed by her.

Her body was taken to Merrifield and was buried on 16 June alongside that of her husband in the Wadham Chapel inside the Church of St Mary, Ilminster, Somerset.