The invention relates to the technical field of biological medicine, and concretely relates to a construction method of a human pancreatic cancer tissue organoid model. The method comprises the following steps: S1, washing a fresh human pancreatic cancer specimen with a cleaning solution, and mechanically cutting into pieces; S2, carrying out enzyme digestion on the mechanically cut tissue fragments; S3, carrying out enzyme-free digestion on the tissue fragments subjected to enzyme digestion, collecting supernatant, and centrifuging to obtain pancreatic cancer single cell precipitate; and S4, fully and uniformly mixing the pancreatic cancer single cells obtained in the step S3 with a matrix material, inoculating gel drops, and adding the gel drops into a complete culture medium for culture. The same pancreatic cancer specimen is sequentially treated by adopting mechanical shearing, enzyme digestion and non-enzyme digestion, and single cells and cell masses with excellent organoid yield are separated, so that the utilization rate of the pancreatic cancer specimen is increased, the quantity and activity of inoculated pancreatic cancer cells are ensured, the number of the organoids meeting the requirement of high-throughput clinical drug screening can be obtained within a short culture time, and a solid foundation is laid for clinical transformation of the organoids.