The invention discloses a bionic three-dimensional DPCs independent co-culture system and a construction method thereof. The construction method comprises the following steps: preparing single-cell suspensions of DPCs, DSCs and HGMCs; providing Matrigel, adding the DPCs single-cell suspension into the Matrigel, and carrying out resuspension to form a Matrigel-DP cell suspension; enabling the Matrigel-DP cell suspension to be adhered to a microporous membrane of a Transwell cell, so as to prepare a three-dimensional DPCs culture system; inoculating HGMCs in an upper chamber of the Transwell cell of the three-dimensional DPCs culture system, simultaneously inoculating DSCs in a lower chamber, and then adding a culture medium to prepare the bionic three-dimensional DPCs independent co-culturesystem. The in-vivo microenvironment of the DPCs can be effectively simulated, the difference between in-vitro cultured DPCs and in-vivo normal DPCs is reduced, and in-vitro long-time maintenance ofbiological functions of the DPCs is realized.