A set of six specific primers were designed based on glycoprotein gene of hirame rhabdovirus(HRV) using Primer Explorer V3 software.The parameter of reaction time and temperature was optimized.The primers were specific with HRV and could not amplify spring viraemia of carp virus(SVCV),infectious pancreatic necrosis virus(IPNV),infectious haematopoietic necrosis virus(IHNV),marine birnavirus(MABV),viral haemorrhagic septicemia virus(VHSV) or viral nervous necrosis virus(VNNV).The detection limit of LAMP assay was approximately 30 fg HRV RNA,which was 100-fold higher than that of conventional RT-PCR.The LAMP reaction could be finished within an hour and clinical fish samples were detected using LAMP.The results showed that 3 fish out of 40 samples were infected with HRV by LAMP assay,which was consistent with virus isolation method.The LAMP assay has great potential in field diagnosis of early HRV infection.
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孙颖杰,岳志芹,刘 荭,赵玉然,史秀杰,梁成珠,吴 斌,李 叶,王 哲. Development and Application of Loop Mediated Isothermal Amplification for Fast Detection of Hirame Rhabdovirus[J]. Jorunal of Huazhong Agricultural University,2010,29(2):203-207.