Development of a gene reporter system in moderately halophilic bacteria by employing the ice nucleation gene of Pseudomonas syringae.

  • Arvanitis N
  • Vargas C
  • Tegos G
 et al. 
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Abstract

The expression of the ice nucleation gene inaZ of Pseudomonas syringae
in several moderate halophiles was investigated to establish its
utility as a reporter for promoter activity and gene expression studies
in these biotechnologically and environmentally important bacteria.
A promoterless version of inaZ was introduced in two different restriction
sites and at both orientations in a recombinant plasmid able to replicate
in moderate halophiles and, in particular, within the sequence of
its pHE1 part, a native plasmid of Halomonas elongata. One orientation
of both recombinant constructs expressed high levels of ice nucleation
activity in H. elongata and Volcaniella eurihalina cells, indicating
that inaZ was probably introduced in the correct orientation downstream
of putative native promoters. A recombinant construct carrying a
tandem duplication of inaZ at the same orientation gave significantly
higher ice nucleation activity, showing that inaZ is appropriate
for gene dosage studies. The ice nucleation gene was also expressed
in H. elongata and V. eurihalina under the control of Pbla (the promoter
of the beta-lactamase gene of Escherichia coli) and Ppdc (the promoter
of the pyruvate decarboxylase gene of Zymomonas mobilis). One of
the inaZ reporter plasmids expressing high levels of ice nucleation
activity under the control of a native putative promoter was also
transferred in Halomonas subglaciescola, Halomonas meridiana, Halomonas
halodurans, and Deleya halophila. In all cases, Ice+ transconjugants
were successfully isolated, demonstrating that inaZ is expressed
in a wide spectrum of moderately halophilic species.

Author-supplied keywords

  • Bacterial Outer Membrane Proteins; Bacterial Prote
  • Bacterial; Genes
  • Genetic
  • Reporter; Genetic Techniques; Gram-Negative Aerob

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  • PMID: 8526492

Authors

  • N Arvanitis

  • C Vargas

  • G Tegos

  • A Perysinakis

  • J J Nieto

  • A Ventosa

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