A fluorescent aptasensor for the Ochratoxin A (OTA) assay, which combines the high efficiency of hyperbranched rolling circle amplification (HRCA) and the specific recognition of the aptamer has been developed. OTA can bind with the aptamer with high affinity, which hinders hybridization between the aptamer and capture probe DNA (CDNA). Then the free CDNA hybridizes with the padlock probe and initiates a HRCA reaction. The HRCA products contain large amounts of double strand DNA (dsDNA), which can then combine with SYBR Green I to produce a fluorescence signal. The fluorescence intensity of the system has a linear relationship with the logarithm of the OTA concentration in the range of 4 fg mL⁻¹ to 400 pg mL⁻¹ and a limit of detection (LOD) of 1.2 fg mL⁻¹ is obtained. The fluorescent aptasensor was applied to detect OTA in corn and oat samples with satisfying results.