The invention discloses a preparation method of gold nano-particle with single-layer protection and relates to a preparation method of gold nano-particle. The invention provides a preparation method of gold nano-meter with mercaptoacetic sodium single-layer protection has the advantages of simple process flow, being environment-friendly, low cost, high rate of finished product and good stability.The composition of the gold nano-meter with mercaptoacetic sodium single-layer protection is gold nano-particle and mercaptoacetic sodium of single-layer protective agent. The hydrochloroauric acid solution with 100 to 300 milligram per liter of gold concentration is prepared; water is added into dry fungal powder R08, and the solution is oscillated to obtain the suspension solution of dry fungalR08 and water; the suspension solution of dry fungal R08 and water is added into the hydrochloroauric acid solution for obtaining the action solution of hydrochloroauric acid and cell R08; The actionsolution of hydrochloroauric acid and cell R08 is oscillated; the mercaptoacetic sodium is added into the action solution of hydrochloroauric acid and cell R08 and oscillated to obtain the mixed solution of gold nano-particle with mercaptoacetic sodium single-layer protection and cell R08; the mixed solution of gold nano-particle with mercaptoacetic sodium single-layer protection and cell R08 is centrifuged to remove the supernatant to obtain the finished product.