Provided is a method of manufacturing a strained silicon-on-insulator (SSOI) substrate that can manufacture an SSOI substrate by separating a bonded substrate using a low temperature heat treatment. The manufacturing method includes: providing a substrate; growing silicon germanium (SiGe) on the substrate to thereby form a SiGe layer; growing silicon (Si) with a lattice constant less than a lattice constant of SiGe on the SiGe layer to thereby form a transformed Si layer; and implanting ions on the surface of the transformed Si layer, wherein, while growing of the SiGe layer, the SiGe layer is doped with impurity at a depth the ions are to be implanted. Accordingly, it is possible to manufacture a substrate with an excellent surface micro-roughness. Since a bonded substrate can be separated using low temperature heat treatment by interaction between implanted ions and impurity, it is possible to reduce manufacturing costs and facilitate an apparatus.