Wilkinson attended school at Harrow and at Exeter College, Oxford. However, he left Oxford in 1818 before earning a degree and joined the army. He loved to travel and made his first visits to the Continent in 1817 and 1818. In 1819, he set off on a tour through France, Germany and Italy, where he met the antiquarian and student of Egyptian hieroglyphics, Sir William Gell. Gell encouraged Wilkinson to leave the army and study Egyptology under his guidance, and in October of 1821, Wilkinson made his first visit to Egypt, at the age of 24, by way of Alexandria, which was the normal entry point in those days. This was a a year before Champollion rediscovered the principles of the Egyptian script. 

This was not a short stay, and his visit marks somewhat of a new beginning for Egyptology. Previously, Egypt had been the focus of brutal exploitation by adventurer such as Drovetti, Henry Salt and Belzoni  His journey to Egypt signals the arrival of men more interested in scientific recording of the monuments in situ, rater than the wholesale removal of objects. He remained in Egypt until 1833, traveling extensively through the country. He learned Cotpic and Arabic, and continued his study of Hieroglyphics. He also managed to survey and record the remains of ancient Egyptian society, including the tombs on the West Bank at Thebes (modern Luxor). In fact, there was hardly an ancient Egyptian site known at that time that Wilkinson did not visit and record in his notebooks. His interest was almost an obsession. There was no inscription, regardless of how small or incomplete that was too insignificant for his attention. 

There, working in 1824 and again between 1827 and 1828, he copied scenes and inscriptions in the private tombs and also surveyed the known tombs in the Valley of the Kings, and produced the first comprehensive plan of ancient Thebes, as well as a chronology of the New Kingdom dynasties. He assigned numbers to the twenty tombs then visible, establishing the numbering system still used today. In fact, modern Egyptologists frequently consult his notes on Thebes particularly for private tomb scenes which were copied by him but since have been damaged or even completely destroyed.

Between 1841 and 1849, he returned to Egypt to survey the Wadi Natron. He spent the winter of 1849-1850 in Italy studying the Turin Canon of Kings, and published a new translation of that composition establishing the correct order of the rulers of ancient Egypt. He traveled to Egypt for the last time in 1855-1856. He worked at the Labyrinth at Hawara and identified it as the mortuary temple of Amenemhat III, and he is also noted as the first individual to map and superficially explore the area of el-Amarna. Others sites he worked at include the tombs at Beni Hasan and at Gebel Barkal. He traveled as far south as the Nubian region. 

Some of his most valuable work was in the field of Epigraphy, which is the study of ancient inscriptions. Through this work, he was able to identify the names of many of the ancient Egyptian kings for the first time. His copies of texts and other drawings are extremely precise, and his notes, now in the Bodleian Library in Oxford remain a mass of information on various aspects of the ancient civilization. In 1837 and 1841, John Murray published a major book by John Wilkinson,  in three volumes, on daily life under the pharaohs called "The Manners and Customs of the Ancient Egyptians, including their private life, government, laws, arts, manufactures, religion, agriculture, and early history, derived from a comparison of the paintings, sculptures, and monuments still existing, with the accounts of ancient authors". The book, which is still used as a reference today, was the first reliable and comprehensive work regarding Egyptian antiquities and remained the best general treatment of ancient Egypt for almost fifty years. A second edition appeared in 1847. This work enhanced Wilkinson's reputation as an Egyptologist, which resulted in his knighthood in 1839 and which made him the first British Egyptologist of distinction. In 1852, he was also granted a DCL from the Oxford University. 

In 1856 Wilkinson married Caroline Catherine Lucas (1822-81), a keen botanist and antiquarian as well as an actress and the companion of Augusta, Lady Llanover. The couple lived first at Tenby in Pembrokeshire, on the South Wales coast. In 1866 they moved to Brynfield House, at Reynoldston on the Gower peninsula. Brynfield and the surrounding area provided Wilkinson with ample opportunity to indulge his interest in ancient British remains. He had already published several articles on British archaeology and antiquities.

Wilkinson's Egyptological work contributed to the foundation of that discipline in Britain, but his research and publications ranged beyond Egypt into architecture, aesthetics, international relations and the classics, as well as travel and the study of ancient Britain. Moreover, in his detailed water-colors and drawings, as in his extensive notes and 'journals', he recorded his impressions of the architecture, costume and contemporary society of all the countries he visited.

On his death in October 19, 1875 at Llandovery in Wales, Wilkinson's library and papers were bequeathed to Sir John Crewe and his family, and were sent to Calke Abbey. It was known that his publications represented only a small proportion of his work, and interest in his papers continued. In 1925 many of the manuscripts relating to Wilkinson's Egyptological research were lent to Francis Llewellyn Griffith, professor of Egyptology at Oxford. After Griffith's death in 1934, these items passed with his library to the Griffith Institute in Oxford. They were used by Bertha Porter and Rosalind Moss in the preparation of their Topographical Bibliography of Ancient Egyptian Hieroglyphic Texts, Reliefs and Paintings (Oxford, 1927-51; revised and reissued, 1960-95), and were given reference numbers used in that work.

In 1984 the National Trust became the owner of Calke Abbey and its contents, including all Wilkinson's manuscripts, which were soon afterwards placed on deposit at the Bodleian, in 56 large volumes (Wilkinson's library remains at Calke.). They were used by Professor Jason Thompson in his biography of Wilkinson (Sir Gardner Wilkinson and his Circle , Austin, Texas, 1992).

After the rise of professional Egyptology in the middle and later nineteenth century, Sir Gardner Wilkinson came to be viewed as an amateur and his popularity diminished. However, recent scholarly work has shown him to be one of the true founders of the discipline. 

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