In this study, we introduce a novel approach to enhance the strength of face-centered cubic (FCC)-structured 316L austenitic stainless steel (ASS) while preserving its tensile ductility. The proposed method involves subjecting the 316L ASS to a pre-strain of 3.5 % at a cryogenic temperature of 4.2 K, resulting in the formation of multiple-fine-defects, including mechanical twins, stacking faults, hexagonal close-packed phases, and body-centered cubic (BCC) phases, which act as BCC martensite seeds. The formation of multiple-fine-defects in 316L ASS through pre-straining at 4.2 K leads to a substantial increase in the yield strength of approximately 45 %, while also maintaining tensile ductility during the subsequent tensile testing at room temperature. Moreover, these pre-existing defects can effectively suppress cross-slip, resulting in a high fraction of planar BCC martensite, which lead to a higher strain-hardening ability than that of the as-annealed 316L ASS. These findings suggest that the mechanical properties of metastable FCC alloys with low to medium stacking fault energies can be effectively enhanced by facilitating martensite seeds via pre-straining at cryogenic temperatures.