Protein expression in Escherichia coli at 15-25 degrees C is widely used to increase the solubility of recombinant proteins. However, many recombinant proteins are insolubly expressed even at those low temperatures. Here, we show that recombinant proteins can be expressed as soluble forms by simply lowering temperature to 6-10 degrees C without cold adapted chaperon systems. By using E. coli Rosetta-gami2(DE3), we obtained 1.8 and 0.9 mg of Ctyptopygus antarticus mannanase (CaMan) and cellulase (CaCel) from 11 culture grown at 6 and 10 degrees C, respectively. Cultivation at 10 degrees C also led to successful expression of EM3L7 (a lipase isolated from a metagenomic library) in a soluble form in E. coil BL21(DE3). Consequently, E. coli cultivation at 6-10 degrees C is an effective strategy for overcoming a major hurdle of the inclusion body formation. (C) 2012 Elsevier Inc. All rights reserved.