[Cloning and expression of TPS1 gene in Escherichia coli]

A S. cerevisiae TPS1 gene for trehalose-6-phosphate synthase was cloned by PCR amplification. The 1.5 kb DNA fragment was ligated to pUC19 and transformed into otsA deficient and deleted of E. coli strains FF4169 and FF4050 separately. otsA gene is encoding trehalose-6-phasphate synthase in E. coli. Restriction endonucleases digestion analysis of transformants' plasmid DNA showed that there was a cloned 1.5 kb fragment carried on the vector. The growth curve experiment result showed that the both transformants could grow well as wild type. Trehalose was synthesized and accumulated in these transformants during high osmotic stress by HPLC combined with ELSD (Evaporate light scatter detector) determination. From the result above, we could conclude that the TPS1 gene of S. cerevisiae was able to restore otsA gene function in E. coli for both osmotolerance and trehalose accumulation during salt stress.