摘要
用TaqMan探针技术设计探针和引物，建立了检测石斑鱼神经坏死病毒（NNV）的实时荧光定量RT-PCR方法.对影响PCR反应的主要因素Mg^2＋浓度和退火温度进行了优化，表明当Mg^2＋浓度为3.0～3.5 mM,退火温度为58～59℃时可获得最佳的扩增和检测效果.应用鞍带石斑鱼（Epinephelus lanceolatus）NNV主衣壳蛋白基因组片段构建的噬菌体假病毒作为阳性质控品,具有很好的稳定性.鞍带石斑鱼NNV的检测试验表明，所建立的实时荧光定量RT-PCR方法是检测石斑鱼神经坏死病毒的一种快速、灵敏、准确的检测方法。
The probe and primers were designed by the way of TaqMan, and the method to examine the nervous necrosis virus （NNV） in reared Epinephelus lanceolatus by the real-time fluorescence quantitative PCR was developed. The main factors of the different concentrations of Mg^2 ＋ and the anneal tempemture affecting the RT-PCR reaction were tested, and it showed that the optimal reaction system was 3.0 to 3.5 mM of Mg^2＋ and 58 to 59 ℃ for anneal temperature. The falsevirus of bacteriophage embodying the major capsid protein of NNV in the tested grouper was treated as the positive contrast, and it displayed the good stability for the detection. It indicated that the method applied in this paper appeared to be quick, sensitive and veracious in detecting the NNV in grouper.
出处
《高技术通讯》
CAS
CSCD
北大核心
2006年第4期431-434,共4页
Chinese High Technology Letters
基金
863计划（2003AA603011）和福建省科技厅科技重大项目（2002N009）资助.
