Patulin (PAT) is a fungal toxin often found in fruit and juice products. The sensitive and fast detection of PAT can help the control of PAT-induced health risk. However, the lack of efficient and stable antibody for PAT hindered the advancement of its test strips. Here, we constructed a nucleic acid-based lateral flow test strip (NALFTS) and used photothermal-colorimetric dual mode for the detection of PAT. The binding of PAT in samples to the aptamer on magnetic separation probe MNPs-Apt@cDNA induced the release of cDNA competitively. Subsequently, cDNA initiates strand displacement amplification (SDA), generating numerous single-strand DNA (ssDNA). These ssDNA molecules were analyzed by a test strip with photothermal-colorimetric dual signal. The developed method demonstrated a limit of detection (LOD) of 5 pg mL−1 and a quantitative range of 0.01 ∼ 320 ng mL−1 in photothermal mode, and a LOD of 0.1 ng mL−1 and a quantitative range of 0.1 ∼ 40 ng mL−1 in colorimetric mode. This SDA-NALFTS offered dual signals corroborating each other to provide more reliable results for rapid screening or accurate monitoring of PAT in juice.