| 对虾IHHNV荧光定量PCR检测方法的建立 |
| Development of a Real - time TaqMan - quantitative PCR Assay for IHHNV Detection |
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| DOI： |
| 中文关键词:对虾 IHHNV病毒 PCR 检测 |
| 英文关键词:Penaeus vannamei IHHNV real - time quantitative PCR detection |
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| 中文摘要: |
| 根据G enBank中对虾传染性皮下和造血器官坏死病毒( IHHNV) 保守基因序列( AF218226), 设计合成了1对引物和1条TaqM an探针, 建立了检测IHHNV的荧光定量PCR技术。将建立的荧光定量PCR检测方法与常规PCR对比。结果显示, 所建立的荧光定量PCR 方法灵敏度可达2个拷贝, 比常规PCR 灵敏度高1 000倍。对保存的15份经常规PCR检测为IHHNV 阳性的DNA 样品进行荧光定量PCR检测, 结果都为阳性, 检测的病毒含量为2. 15 @ 107 ~ 4. 21 @ 104 拷贝/LL。用该方法对不同浓度的样品进行了重复检测, 表明该方法具有良好的重复性, 可满足IHHNV的临床诊断需要。
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| 英文摘要: |
| A pair of primers and a TaqMan probe were designed and synthesized according to the conserved gene sequences of Infectious Hypodermal and Haematopoietic Necrosis （IHHNV） in GenBank （AF218226）, and then reaction parameters were optimized to develop a real -time TaqMan -quantitative PCR assay. The developed quantitative PCR assay was compared with that of routine PCR. This quantitative PCR assay could detect 2 template copies of plasmid DNA, and its sensitivity was 1 000 times higher than that of the routine PCR. The real - time Taq- Man - quantitative PCR results of 15 routine PCR positive clinical samples showed that concentration of the clinical samples were 2.15 ×10^7 -4.21 × 10^4 copies/μL. The samples were examined using the quantitative PCR repeatedly and the results indicated that the quantitative PCR was reproducible and could be used successfully for the diagnosis of IHHNV infection. |
| 谢丽基,谢芝勋,庞耀珊,卢兆发,谢志勤,刘加波,邓显文,唐小飞.2008.对虾IHHNV荧光定量PCR检测方法的建立[J].水生态学杂志,29(2):132-135. |
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