Abstract: The strain product heat resistance protease was screened on the milk agar plate through the separation and purification from the high-temperature daqu samples, then the strain A1 which can produce more enzyme from further purification was selected by measuring the liquid fermentation enzyme protease.The strain A1 was identified as Bacillus thuringiensis through morphological characteristics, biochemical identification, 16S rDNA sequencing and molecular biology.Three main factors which have affected the choice of glucose, yeast extract and pH, was optimized for fermentation medium with response surface method.The optimum medium for enzyme production was pH value of 8.4, glucose dosage 1.092%, yeast extract dosage of 0.902%, the best enzyme activity was 142.81u/mL.Determination of protease enzyme optimum temperature was 61℃, and it was higher enzyme activity when 55 ～ 70℃, the relative activity was 97.63% at 60℃.