The invention discloses a crossbreeding method in a dendrobium test tube. According to the invention, the crossbreeding method in the dendrobium test tube is adopted, the ripening period of fruits can be shortened greatly, and crossbreeding generations in the test tube can bloom within a short period of time, and flower patterns and colors can be observed, and novel varieties are bred, thereby accelerating the dendrobium breeding speed, and the crossbreeding method is reported for the first time internationally; because dendrobium in the test tube can bloom annually, and can crossbreed with varieties rare in flower seasons in the natural world. Additionally, according to the invention, culture medium in each stage is utilized, 'hyponex' is utilized as the main component, and the cultural media are unique in ingredient, not only are low in cost, but also are high in abloom rate, and the fruits are developed rapidly; moreover, only a simple plant tissue culture device is required to implement the crossbreeding method provided by the invention, and therefore the whole breeding method is simple and convenient, is low in cost, and creates conditions for culturing the high-quality dendrobium variety.