A screening study was made in order to evaluate the infection and identification of different fungal genera associated with storage wheat varieties. Total loss due to seed-borne diseases is up to an extent of 30-75%. Disease free quality seed production in wheat is utmost important to sustain the production and to maintain the quality crop. Keeping this in view, detailed investigation were carried out to study the implication of seed-borne fungi of wheat on seed quality parameter and to find out suitable detection methods for seed-borne infection of the three detection methods adopted for inoculation of seed mycoflora in wheat varieties. Wheat varieties grown during the months of October to March 2011, collected from Quarsi agricultural farm, Aligarh were screened by using Blotter method, Agar plate method and Deep freeze method as recommended by ISTA. All experiments were in a completely randomized design (CRD) in three replicates. Out of the five varieties tested, PBW343 was found to be most susceptible and was associated with more fungal flora than the other varieties. Seed mycoflora of abnormal seeds was also studied. Maximum incidence of fungi was observed in case of discolored seeds followed by shrunken seeds and cracked seeds. In all the three detection methods, a total no of 11 genera and 20 species of fungi were isolated and identified .