Extended-spectrum β-lactamases (ESBLs) are enzymes that hydrolyze the β-lactam ring, and ESBL-producing E. coli has rapidly spread worldwide with pose a serious hazard for humans. The aim of this study was to determine the prevalence of ESBL producing E. coli and molecular evaluation of four ESBL-associated genes among E. coli strains isolated from milk and cheese in southern Iran. Antibiotic susceptibility test was carried out for a total of 150 isolates of E. coli, previously collected from dairy products. ESBL production was screened using a double-disc synergy test (DDST) and presence of four ESBL genes (PER, VEB, TEM and CTX-M) was tested using PCR. Among 150 E. coli strains 57 (38%) isolates were identified as ESBL-producing strains. All ESBL positive isolates could be typed for one or more genes and the most prevalent ESBL-associated gene was CTX-M (80.7%). The PER gene was not present among isolates. Isolates showed high susceptibility to imipe¬nem and cefoxitin. The results showed the high prevalence of ESBL producing E. coli strains among dairy products and high occurrence of CTX-M-associated ESBL activity among isolates indicating the hazards of increasing the strains with antibiotic resistance which can transfer to human trough the dairy food products.