Publication: Construction and characterization of regulated L-arabinose-inducible broad host range expression vectors in Xanthomonas
Issued Date
1999-12-15
Resource Type
ISSN
03781097
Other identifier(s)
2-s2.0-0032709116
Rights
Mahidol University
Rights Holder(s)
SCOPUS
Bibliographic Citation
FEMS Microbiology Letters. Vol.181, No.2 (1999), 217-223
Suggested Citation
Rojana Sukchawalit, Paiboon Vattanaviboon, Ratiboot Sallabhan, Skorn Mongkolsuk Construction and characterization of regulated L-arabinose-inducible broad host range expression vectors in Xanthomonas. FEMS Microbiology Letters. Vol.181, No.2 (1999), 217-223. doi:10.1016/S0378-1097(99)00541-8 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/25321
Research Projects
Organizational Units
Authors
Journal Issue
Thesis
Title
Construction and characterization of regulated L-arabinose-inducible broad host range expression vectors in Xanthomonas
Other Contributor(s)
Abstract
Several versions of broad host range (BHR), L-arabinose-inducible expression vectors were constructed. These expression vectors were based on a high copy number BHR pBBR1MCS-4 replicon that could replicate in both enteric and non-enteric Gram-negative bacteria. Two versions of expression cassettes containing multiple cloning sites either with or without a ribosome binding site were placed under transcriptional control of the Escherichia coli BAD promoter and araC gene. Three versions of vectors containing ampicillin or kanamycin or tetracycline resistance genes as selectable markers were constructed. In all six new L-arabinose-inducible BHR expression vectors containing many unique cloning sites, selectable markers were made to facilitate cloning and expression of genes in various Gram-negative bacteria. A Tn9 chloramphenicol acetyl transferase (cat) gene was cloned into an expression vector, resulting in pBBad18Acat that was used to establish optimal expression conditions (addition of 0.02% L-arabinose to mid-exponential phase cells for at least 1 h) in a Xanthomonas campestris pv. phaseoli. Comparison of the Cat enzyme activities between uninduced and a 180-min L-arabinose-induced culture showed a greater than 150-fold increased Cat specific activity. In addition, L-arabinose induction of exponential phase cells harboring pBBad18Acat gave a higher amount of Cat than similarly treated stationary phase cells. The usefulness of the expression vector was also demonstrated in both enteric and non-enteric Gram-negative bacteria. Copyright (C) 1999 Federation of European Microbiological Societies.