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Species‐specific polymerase chain reaction primers for Lactococcus garvieae
Author(s) -
Aoki T.,
Park CI.,
Yamashita H.,
Hirono I.
Publication year - 2000
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.1365-2761.2000.00207.x
Subject(s) - biology , polymerase chain reaction , agarose gel electrophoresis , dna extraction , primer (cosmetics) , enterococcus faecalis , microbiology and biotechnology , enterococcus faecium , enterococcus , lactococcus , dna , gene , bacteria , lactococcus lactis , genetics , escherichia coli , chemistry , antibiotics , lactic acid , organic chemistry
A dihydropteroate synthase gene from the chromosomal DNA of the fish‐pathogenic bacteria Lactococcus garvieae (formerly Enterococcus seriolicida ) was cloned. This gene was then chosen as the target for polymerase chain reaction (PCR). The designated PCR primer set only amplified a 709‐bp DNA fragment from L. garvieae strains, and did not amplify the same molecular size fragment from related species of L. lactis, Enterococcus faecalis, E. faecium or β‐haemolytic Streptococcus sp. The kidney tissue of yellowtail, Seriola quinqueradiata (Temminck and Schlegel), a species which is naturally infected with L. garvieae , and also kidney tissue samples of healthy yellowtail were stored in TNES‐Urea. The DNA was extracted from tissue samples by a modification of the standard method and by a boiled‐extraction method. In particular, template DNA was utilized within 30 min following extraction and purification by the boiled‐extraction method. These species‐specific PCR primers could amplify a L. garvieae target sequence from yellowtail which were naturally infected with L. garvieae . The total procedure for the diagnosis of L. garvieae infections in fish, from the point of DNA extraction to observation in an agarose gel following electrophoresis, can be performed in less than 4 h.