Strain GIHVLr6426 was isolated as an isoleucine hydroxamate-resistant mutant from the wild-type strain of Serratia marcescens Sr41. This mutant had high activities of threonine deaminase and acetohydroxyacid synthase, both of which were insensitive to feedback inhibition. Genetic analysis revealed that strain GIHVLr6426 carried two regulatory mutations located in the ilv region. Strain T-693, a threonine-producing strain which was previously reported to carry four regulatory mutations for threonine biosynthesis, had an increased activity of acetohydroxyacid synthase. Transductional analysis revealed that one of the four mutations carried by strain T-693 was responsible for constitutive synthesis of both isoleucine and threonine biosynthetic enzymes. Strain GIHVLr6426 into strain T-693. The constructed strain had the six regulatory mutations for threonine and isoleucine biosyntheses, and produced about 25 mg isoleucine ml-1 in a medium containing sucrose and urea.