| EHP、SHIV双重TaqMan实时荧光定量PCR检测方法的构建及应用 |
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| 引用本文： | 侯月娥,曾俊霞,蓝间媛,许枣珠,廖秀云,罗宝正.EHP、SHIV双重TaqMan实时荧光定量PCR检测方法的构建及应用[J].中国动物传染病学报,2022(1):112-119. |
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| 作者姓名： | 侯月娥 曾俊霞 蓝间媛 许枣珠 廖秀云 罗宝正 |
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| 作者单位： | 珠海科艺普检测科技有限公司;拱北海关技术中心 |
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| 基金项目： | 珠海市公共技术服务平台产学研协同创新计划项目(IETP201901011)。 |
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| 摘 要： | 本研究根据GenBank中已有的虾肝肠胞虫(EHP)和虾血细胞虹彩病毒(SHIV)基因的保守序列,设计特异的引物和探针.建立了快速诊断EHP和SHIV的双重TaqMan实时荧光定量PCR检测方法,并对其特异性、敏感性和稳定性进行检测.结果表明:该方法检测限可达10 copies/μL,其敏感性是SYBR Green r...
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| 关 键 词： | 虾肝肠胞虫 虾血细胞虹彩病毒 TaqMan荧光定量PCR |
Development of a Duplex TaqMan Real-time PCR Assay for Detection of Enterocy Tozoon Hepatopenaei and Shrimp Hemocyte Iridovirus |
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| HOUYuee,ZENG Junxia,LAN Jianyuan,XU Zaozhu,LIAO Xiuyun,LUO Baozheng.Development of a Duplex TaqMan Real-time PCR Assay for Detection of Enterocy Tozoon Hepatopenaei and Shrimp Hemocyte Iridovirus[J].Chinese Journal of Animal Infectious Diseases,2022(1):112-119. |
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| Authors: | HOUYuee ZENG Junxia LAN Jianyuan XU Zaozhu LIAO Xiuyun LUO Baozheng |
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| Institution: | (Zhuhai Kerric Testing Technolongy Co.Ltd.,Zhuhai 519000,China;Gongbei Customs district technology center,Zhuhai 519015,China) |
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| Abstract: | In this study,specific probes and primers were designed according to Enterocy Tozoon Hepatopenaei(EHP)and Shrimp Hemocyte Iridovirus(SHIV)gene sequences in GenBank.A duplex TaqMan-based real-time quantitative PCR assay was then developed by optimizing the reaction conditions.The results demonstrated that this PCR assay was more sensitive for EHP and SHIV with the detection limit of 1×10 copies/μL,which was 10-fold higher than SYBR Green real-time PCR and 100-fold higher than conventional PCR.Specifi city tests showed no cross reactions with White spot syndrome virus,Infectious hypodermal and hematopoietic necrosis virus,High pathogenicity ibrio parahaemolyticus and Taura syndrome virus.Repeatability tests demonstrated that the threshold cycle of the method gave a good stability with the coeffi cient of variations were less than 4%.Then 37 samples were tested by using a duplex TaqMan real-time PCR and conventional PCR and the results reached 100%agreement.In conclusion,the duplex TaqMan-based real-time quantitative PCR assay developed here were specifi c and sensitive for rapid detection of EHP and SHIV.Therefore,it could be suitable for early duplex TaqMan-based real-time quantitative PCR assay of latent infection of shrimp. |
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| Keywords: | Enterocy tozoon hepatopenaei shrimp hemocyte iridovirus TaqMan real-time PCR detection |
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