Abstract: Fumonisin B₁ (FB₁) is a water-soluble mycotoxin produced mainly by Fusarium during the growth of agricultural products. FB₁ is mostly distributed in grains and their products represented by corn. It can cause acute poisoning of livestock, damage the immune system of animals, and has potential carcinogenicity. Therefore, it is of great significance to improve the detection level of the contamination of FB₁ in the food matrix for reducing economic losses and ensuring public health. In this work, the highly specific new recognition probe-aptamer and the easy-to-operate fluorescence analysis method were combined and applied. Using nanomaterials and RecJ_f exonuclease as signal sensing strategies, a simple and stable fluorescence sensor with high sensitivity was developed for FB₁ detection. Nitrogen doped graphenes (NDGRs) with large specific surface area and good dispersion were prepared and used as the substrate of the aptamer recognition probe of FB₁. NDGRs can effectively quench the fluorescence of carboxyfluorescence (FAM) labeled FB₁ specific aptamer due to their strong π-π stacking interaction. With the addition of FB₁, it could bind with aptamer to form FAM-Aptamer/FB₁ complex, which cannot adsorb on the surface of NDGRs and thus retains the dye fluorescence. Meanwhile, the FAM-Aptamer/FB₁ complex can be cleaved by RecJ_f exonuclease, and in such case FB₁ is delivered from the complex. The released FB₁ then binds with other FAM-labeled aptamers on the NDGRs surface, and touches off another target recycling, resulting in the successive release of dye-labeled aptamers from the NDGRs, which leads to a significant amplification of the signal. The optimized experimental conditions were as follows:the concentration of NDGRs was 40 μg/mL, the optimal amount of RecJ_f exonuclease was 5 U and the incubation time was 90 min. Under the experimental conditions, the standard curve of the aptasensor was established. The fluorescence intensity of the proposed aptasensor had a good linear with FB₁ concentration in the range of 0.2-20 ng/mL and 20-500 ng/mL, respectively. The detection limit was 0.084 ng/mL. The experimental results demonstrated that the aptasensor showed good specificity toward FB₁ in the anti-interference performance evaluation results for AFB₁, OTA and ZEN. Meanwhile, the corn and beer spiked samples were tested and the obtained average recovery rates were 91%-103% and 92%-97%, respectively, indicating that the prepared sensor is suitable for the detection of FB₁ in food. The prepared aptasensor is of great practical importance in food safety and can be widely extended to the detection of other toxins by replacing the sequence of the recognition aptamer.