Cloning and analysis of promoter-active fragments from Corynebacterium glutamicum 10147

Abstract:

Abstract: [Objective] To clone promoter-active fragments from Corynebacterium glutamicum for further construction of expression vectors. [Methods] Random Sau3A I digested fragments of C. glutamicum 10147 chromosome were shot-gun cloned into the promoter-probe vector pAKC6 and promoter activity of the inserted fragments was selected by chloramphenicol resistance of transformed C. glutamicum cells. [Results] Thirty promoter-carrying fragments were isolated. Three C. glutamicum clones harboring pAKC6 with promoter fragments displayed chloramphenicol acetyltransferase (CAT) activity of more than 24 U/mg. The fragment F57 led to the highest CAT activity of 32.50 U/mg, even more than that produced by the promoter Ptrc, 26.33 U/mg. [Conclusion] The strength of promoter on fragments F21, F54 and F57 is as strong as promoter Ptrc in C. glutamicum. These fragments can be used to construct expression vector.

Key words:Keywords: Corynebacterium glutamicum; promoter-probe vector; promoter; chloramphenicol acetyltransferase