Philippine Science Letters
vol. 4 | no. 2 | 2011
published online December 9, 2011


ARTICLE


Amplification and sequence analysis of the luxA gene in the non-luminescent type strain Vibrio campbellii NBRC 15631


by Cynthia T. Hedreyda* and Fabini D. Orata


National Institute of Molecular Biology and Biotechnology, College of Science, University of the Philippines, Diliman, Quezon City, Philippines 1101


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Type strains of V. campbellii can be distinguished from its close relative V. harveyi by luminescence activity where V. campbellii tests negative and V. harveyi tests positive. The luminescent phenotype depends on the ability to produce the enzyme luciferase that has two subunits, alpha (a) and beta (▀), encoded by adjacent genes luxA and luxB, respectively. The luxA and luxB genes have been isolated and sequenced from type strain V. harveyi, but only the luxB gene has been amplified and sequenced from type strain V. campbellii. In this study, the luxA gene from V. campbellii NBRC 15631 was amplified and sequenced in order to gain insight into the molecular basis of the non-luminescent phenotype in this Vibrio. Inverse PCR using circularized HindIII-digested genomic DNA of type strain V. campbellii as template and primers that amplify genes flanking the luxB gene produced a 1,100 bp amplicon containing the 225 bp luxA gene. This much shorter luxA of V. campbellii NBRC 15631 and the full length 1,068 bp type strain V. harveyi NBRC 15634 luxA showed only 88% sequence similarity. The 225 bp luxA of V. campbellii corresponds to a 74-amino acid LuxA polypeptide exhibiting only 74% sequence similarity with LuxA of type strain V. harveyi. The much shorter V. campbellii luxA is expected to give rise to a variant LuxA polypeptide that may have produced a luciferase with an altered and non-functional active site. This study, therefore, provides a molecular basis for the difference in luminescence phenotype between the type strains of the two closely related species, V. harveyi and V. campbellii. Moreover, the significant difference in the luxA gene sequence between type strains of V. harveyi and V. campbellii can be utilized in the detection and identification of these two closely related species.


Email Address: chedreyda@mbb.upd.edu.ph
Submitted: May 6, 2011
Revised: July 25, 2011
Accepted: October 1, 2011
Published: December 9, 2011
Editor-in-charge: Manuel M. Garcia
Reviewers:
Olga Andrievskaia
Hongsheng Huang