The materials used in this research is ‘Benihoppe’, a popular cultivar in strawberry production with weak resistance to black rot disease. We isolated and identified the pathogen of black rot disease from sick strawberry plants. Then, we further studied on the effect and mechanism of exogenous melatonin on improving the resistance of strawberry to black rot. We detected the pathogenicity of strawberry black rot, the bacteriostatic effect, the antibacterial ability of melatonin on strawberry, the changes of related enzymes activities and expression level of resistancerelated genes in the leaves with spraying exogenous melatonin and inoculating the pathogen of strawberry black rot. The results showed that: (1) the separation, sequence analysis and infection test proved that the pathogen was Alternaria alternate, which can result in black rot disease in ‘Benihoppe’ strawberry. (2) The hyphae growth rate of A. alternate was inhibited to different degrees on PDA media containing different concentrations of melatonin 1, 2, 4 and 8 mmol/L. It had slower growth rate with higher melatonin concentration. The inhibition rate of hyphae of A. alternate on the medium containing 8 mmol/L melatonin was 68.9%. (3) We found that exogenous melatonin had the function of delaying the infected onset process of pathogen (A. alternate) in leaves and stolons after they were pretreated for 24 hours. The inhibitory effect increased with the increase of melatonin concentration and 0.5 mmol/L melatonin was the most effective in inhibiting pathogen infection. (4) Biochemical analysis showed that 0.5 mmol/L exogenous melatonin pretreatment could improve the activities of diseaserelated enzymes such as CAT, POD, PAL and PPO after the strawberry leaves infected with pathogen, among which the activity of PPO changed most significantly, with a 23.8% higher inhibition rate than that of the control group. (5) 0.5 mmol/L exogenous melatonin treatment could significantly increase the expression levels of diseaserelated genes such as PR1Alike, PR10, WRKY1, PPO and CCR in strawberry. It suggested that exogenous melatonin could effectively inhibit the hyphae growth rate of A. alternate and delay the onset process. The best concentration was 0.5 mmol/L. And WRKY1 transcription factor plays an important regulatory role during the process of improving strawberry resistance to black rot disease. Exogenous melatonin may improve strawberry resistance to black rot by activating the expression of this transcription factor to regulate the expression of related genes and the activities of related antioxidant enzymes and defense enzymes.