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Molecular cloning of Yersinia ruckeri aroA gene: a useful taxonomic tool
Author(s) -
Yugueros J,
Temprano A,
Luengo J M,
Naharro G
Publication year - 2001
Publication title -
journal of fish diseases
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.819
H-Index - 85
eISSN - 1365-2761
pISSN - 0140-7775
DOI - 10.1046/j.1365-2761.2001.00311.x
Subject(s) - aroa , biology , yersinia ruckeri , polymerase chain reaction , microbiology and biotechnology , gene , plasmid , nucleic acid sequence , molecular cloning , genetics , escherichia coli , complementary dna , enterobacteriaceae , fishery , fish <actinopterygii> , rainbow trout
The aroA gene of Yersinia ruckeri , which encodes 5‐enolpyruvylshikimate 3‐phosphate (EPSP) synthase was cloned by complementation of the aroA mutation in Escherichia coli AB2829 by using pUC18 plasmid as a vector. Nucleotide sequence of the aroA gene revealed an open reading frame of 427 amino acids showing a high degree of homology to other bacterial AroA proteins. A pair of primers with 23 and 20 nucleotides were selected from the 5′ and 3′ termini, respectively, and formed the basis of a specific polymerase chain reaction (PCR) assay. A 1165‐bp deoxyribonucleic acid (DNA) fragment was amplified from all lysed Y. ruckeri strains. An identical size fragment was also amplified from lysed Y. pseudotuberculosis , Y. aldovae , Salmonella enteritidis and E. coli , but not from other enterobacteria. Alu I restriction fragment length polymorphism (RFLP) of the PCR amplified products allowed for differentiation between Y. ruckeri and the other bacteria. Specificity and sensitivity make this PCR assay a useful method for rapid identification and diagnosis of Y. ruckeri infections.