The quadrate (incus) bone underwent important evolutionary transformations through the cynodont-mammal transition. The following character transformations played crucial roles in modifying the cynodont quadrate into the mammalian incus: 1) progressively greater rotation of the dorsal plate relative to the trochlea; 2) the contact facet of the dorsal plate becomes concave; 3) development of a constricted neck between the dorsal plate and the trochlea; 4) simplification of the quadrate-cranium joint, resulting in better mobility of the joint; and 5) introduction of a stapedial process (crus longum). The dorsal plate rotation, the concave contact facet, the constricted neck, the mobile joint of the quadrate and the cranium are also present in some advanced non-mammalian cynodonts. Broad phylogenetic distributions of these features suggest that the major features of the incus of early mammals, as represented by Morganucodon, originated much earlier in phylogenetic history among non-mammalian cynodonts. Apomorphies of the quadrate (incus) among the advanced non-mammalian cynodonts favor a sister-group relationship of tritheledontids and mammals. The hypothesis on the postdentary origin of the mammalian tympanic membrane is favored by transformation of the quadrate through the cynodont-mammal transition. Three most important modifications of the quadrate (incus) through the cynodont-mammal transition are: formation of the concave contact facet, progressively greater rotation of the dorsal plate, and decrease in the number of cranial bones articulating with the quadrate. These modifications would simplify the quadrate-cranial joint and increase the mobility of the quadrate (incus) relative to the cranium while a functioning tympanic membrane was maintained on the mandible, improving the sensitivity of the postdentary tympanum. Probainognathus is among the earliest known non-mammalian cynodonts with a concave contact facet and a rotated dorsal plate in the quadrate. Thus we hypothesize that it represents a critical step in the phylogenetic transformation that led to the origin of the modern mammalian middle ear and tympanic membrane.