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Development of a multiplex polymerase chain reaction to detect five common Gram‐negative bacteria of aquatic animals
Author(s) -
Tsai MA,
Ho PY,
Wang PC,
E YJ,
Liaw LL,
Chen SC
Publication year - 2012
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 - 10.1111/j.1365-2761.2012.01372.x
Subject(s) - biology , aeromonas hydrophila , amplicon , vibrio vulnificus , microbiology and biotechnology , vibrio parahaemolyticus , multiplex polymerase chain reaction , polymerase chain reaction , edwardsiella tarda , arcobacter , genomic dna , bacteria , multiplex , dna , gene , genetics , 16s ribosomal rna
A multiplex polymerase chain reaction (m‐PCR) technique was developed as a rapid and accurate diagnostic tool for identifying five major Gram‐negative bacilli – Vibrio vulnificus , V. parahaemolyticus , Aeromonas hydrophila , Chryseobacterium meningosepticum and Edwardsiella tarda – that cause major diseases in cultured aquatic animals in Taiwan. The expected amplicons for V. vulnificus , V. parahaemolyticus , A. hydrophila , C. meningosepticum and E. tarda were 410, 368, 685, 180 and 230 bp, respectively. The assay was shown to be specific for the target pathogens. The sensitivities of detection were estimated to be 20.5 fg∼200 pg of genomic DNA or 10 2 ∼10 4 colony‐forming units (cfu) of bacterial isolates when adopted as PCR templates. The m‐PCR was capable of simultaneously amplifying target fragments from bacterial genome DNA mixed with the DNA extracted from viscera and tissues taken from fish without affecting the performance of the method.