The invention discloses an expression vector for synthesizing cannabidiol, a heterologous expression method and application, five gene expression cassette vectors are constructed through an in-vitro de novo design and five enzymes of hexanoyl-CoA synthase, polyketide synthase, oleinic acid synthase, isopentenyl transferase and cannabidiol acid synthase, and a transient co-transformation experiment in tobacco shows that in this way, high yield of CBD can be achieved. According to the invention, efficient production of CBD can be realized by using a synthetic biological technical means, the synthetic product does not contain THC, and the measured CBD content is high; in addition, as a conventional commercial crop for agricultural production, the tobacco has advantages in planting technology, field management, yield of harvested leaves and the like, and planting cost can be greatly reduced.