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The present study is concerned with the production of glucose oxidase (GOD) from thermophilic Penicillium sp. in 250 mL shake flask. Fourteen different strains of thermophilic Penicillium sp. were isolated from the soil and were screened for glucose oxidase production. IIBP-13 strain gave maximum extra-cellular glucose oxidase production as compared to other isolates. Effect of submerged fermentation in shaking and static conditions, different carbon sources and incubation period on the production of extra-cellular glucose oxidase was studied. Maximum yield (0.325 U/mL) of extra-cellular glucose oxidase was obtained after 72 hrs of incubation at 45 oC using submerged fermentation technique in shaking conditions utilizing sucrose as carbon source.