The permeability of low-permeability cores is generally measured using a pulse decay method. The core of low-permeability rocks, such as shale, often has a layered structure. The applicability of pulse decay testing for layered cores is not clear. In this study, the performance of the pulse decay method on layered cores was comprehensively investigated. Numerical simulations were conducted to investigate the influence of the interlayer permeability ratio, storativity ratio, layer thickness, interlayer location, and number of layers on the pulse decay pressure and pressure derivative curves, as well as the permeability obtained from pulse decay testing. The results revealed that the pressure curves of layered cores exhibit distinct differences from those of homogeneous cores if the upstream permeability is larger than the downstream one. The pressure derivative curve shows more inclined or horizontal straight-line segments than in the homogeneous case. The shapes of the pressure and pressure derivative curves are affected by the upstream and downstream positions of the core, but the tested permeability is not affected. The tested permeability differs from the equivalent model permeability, with an error of up to 22%. If the number of layers is not less than 10, the permeability obtained from the pulse decay test is consistent with that of the equivalent model. These differences are influenced by the interlayer permeability ratio, storativity ratio, layer thickness, interlayer location, and number of layers. To improve the accuracy of permeability analysis in pulse decay testing for layered cores, curve fitting using the characteristics of the pressure derivative curve can be employed.