The temporary fasteners are used in pre-joining during aircraft assembly. To improve the fit effect of assembly parts in pre-joining process, a layout optimization method for temporary fasteners in composite structure assembly pre-joining was proposed. An equivalent model for calculating the deformation and gaps of the panel in pre-joining process was established, and the calculation of key variables was improved. Layout optimization of temporary fasteners was performed based on the equivalent model. Assembly experiments of composite panel and metal frame wing box was conducted for verification. The results show that the improved calculation method can calculate key matrix variables in a shorter time. The single computation time for panel deformation during the optimization process is significantly reduced. The calculated deformation and gaps results are in good consistency with the experimental data. The established method can calculate the layout of temporary fasteners with high surface fit ratio, providing valuable reference for the development of pre-joining processes for large composite structure assemblies.