The invention relates to a ZnO quantum dot vector / DNA composite-containing collagen-based composite cornea substitute, and a preparation method and application thereof. The ZnO quantum dot vector / DNA composite-containing collagen-based composite cornea substitute is prepared by absorbing ZnO quantum dot vectors / DNA composites by using a collagen / MPDSAH IPN cornea substitute, wherein the weight ratio of the cornea substitute to the ZnO quantum dot vectors / DNA composites is 425:1; and the weight ratio of ZnO quantum dot vectors to DNA is 25:1. The ZnO quantum dot vector / DNA composite-containing collagen-based composite cornea substitute has high biocompatibility, can induce and promote the regeneration of a cornea and be biologically decomposed along with the regeneration of the cornea, hasmechanical properties and optical properties the same as those of a human cornea, remarkably improves the stability and mechanical strength of the cornea substitute in collagenase due to the introduction of an MPDSAH polymer network, can effectively compositely compress the DNA, successfully induce the DNA into cells, successfully express the DNA, trace the position of the DNA / vector at any time and determine the intra-cellular distribution of the DNA / vector in a transgenosis process due to ZnO quantum dots, and effectively combines gene transfection, tissue engineering and fluorescent tracing so as to achieve simple manufacturing method, and easy processing and long-term storage and transport.