The macroscopic property of optical material depends on the chemical composition as well as its mesoscopic structure. Thus, the processing of micro-and nanostructures of soft matter, including liquid crystal, is of great importance and can be exploited in many areas. The laser-assisted mechanical injection brings the concept of laser micro-nano manufacturing to the field of liquid crystal (LC) and injects a certain number of microdroplets (MDs) of the water solution with a uniform size into a host LC drop by using a laser beam. With the help of the elasticity of the LC medium, the injected water MDs spontaneously self-assemble into colloidal crystals as inner structures within the closed 3D space of the host LC drop in the microscale. The self-assembly structure of injected MDs in 3D has been scanned using the confocal microscope. In addition, by introducing the cholesteric LC drops with a topological defect line as the host, the injected MDs are captured by the defect line and self-assembled along its geometry into a chain. Moreover, it is proved that laser-assisted mechanical injection is also available for the interface of two immiscible fluids with a relatively high viscosity. The results provide a new method for fabricating and developing customized hierarchical microemulsion, drug delivery system, biosensor, and functionalized microstructure of soft matter.