Rapid diagnosis of turbot reddish body iridovirus in turbot using the loop-mediated isothermal amplification method.
Turbot reddish body iridovirus (TRBIV) is a new piscine iridovirus that infects the turbot, Scophthalmus maximus, cultured in northern China and can cause high mortality. In this study, a loop-mediated isothermal amplification (LAMP) method was developed for the specific detection of this virus using primers designed from an Msp I restriction DNA fragment of the TRBIV genome. Mg(2+) concentrations, the reaction temperature, and the reaction time of LAMP were optimized to 6mM, 65 degrees C, and 60 min, respectively. The detection limit of the LAMP method was as low as seven copies and was 100 times more sensitive than the conventional PCR technique. Visual inspection of LAMP amplifications demonstrated that the positive and negative reactions exhibit distinct and different colors in daylight, which means that gel electrophoresis is not necessary to judge the presence or absence of the virus. LAMP can be conducted in 1h and requires only a simple heating device for incubation. Thus, the LAMP-TRBIV detection protocol has great potential for use in the detection of TRBIV in both the laboratory and the farm.
Zhang
Qingli
Q
Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao, Shandong, China.
Shi
Chenyin
C
Huang
Jie
J
Jia
Kuntong
K
Chen
Xinhua
X
Liu
Hong
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002681:China; D017931:DNA Primers / Q000235:genetics; D004266:DNA Virus Infections / Q000662:veterinary* / Q000821:virology; D020536:Enzyme Activators / Q000494:pharmacology; D005393:Fish Diseases / Q000175:diagnosis* / Q000821:virology; D005412:Flatfishes / Q000821:virology*; D007497:Iridoviridae / Q000235:genetics / Q000302:isolation & purification*; D008274:Magnesium / Q000494:pharmacology; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D013696:Temperature; D013997:Time Factors
D017931:DNA Primers; D020536:Enzyme Activators; D008274:Magnesium