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J Virol Methods. 2014 Feb;196:199-203. doi: 10.1016/j.jviromet.2013.11.011. Epub 2013 Nov 27.

The development of a loop-mediated isothermal amplification assay for rapid and sensitive detection of abalone herpesvirus DNA.

Author information

1
Institute of Veterinary Medicine, School of Veterinary Medicine, National Taiwan University, Taipei, Taiwan; Tzu Chi College of Technology, Hualien, Taiwan.
2
National Institute for Animal Health, Tansui, Taiwan.
3
Ifremer, Unité Santé Génétique et Microbiologie des Mollusques, Laboratoire de Génétique et Pathologie des Mollusques Marins, 17390 La Tremblade, France.
4
Institute of Veterinary Medicine, School of Veterinary Medicine, National Taiwan University, Taipei, Taiwan. Electronic address: penheng@ntu.edu.tw.

Abstract

A loop-mediated isothermal amplification (LAMP) assay was developed for the detection of abalone herpesvirus DNA. Two pairs of primers were designed, based on the sequence of the DNA polymerase gene of abalone herpesvirus. The reaction temperature and time were optimized to 63°C and 60min, respectively. LAMP amplicons were analyzed by 2% agarose gel electrophoresis or by visual inspection of a colour change emitted by fluorescent dye. The method developed was specific for the detection of abalone herpesvirus, without cross-reactions with other tested herpesviruses including ostreid herpesvirus 1 (OsHV-1), European eel herpesvirus, koi herpesvirus (KHV) and an avian herpesvirus. The LAMP assay was 100 folds more sensitive than a conventional PCR and 10 folds less sensitive than a SYBR Green PCR. These results indicate that the developed LAMP assay is a simple, rapid, sensitive, specific and reliable technique for the detection of abalone herpesvirus.

KEYWORDS:

Abalone; Haliotis diversicolor supertexta; Herpesvirus; Loop-mediated isothermal amplification; Polymerase chain reaction; SYBR green PCR

PMID:
24291740
DOI:
10.1016/j.jviromet.2013.11.011
[Indexed for MEDLINE]
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