传染性造血器官坏死病毒液相芯片检测技术

Development of a Liquid Chip Assay for the Detection of Infectious Haematopoietic Necrosis Virus(IHNV)

用DNAStar7.0软件对GenBank中IHNV的N基因进行序列分析,设计IHNV特异性引物并标记生物素,将探针氨基化修饰,与荧光编码微球偶联后与RT-PCR产物杂交反应,用液相芯片仪器检测荧光信号,建立了传染性造血器官坏死病毒(IHNV)的液相芯片快速检测技术。结果表明液相芯片检测体系对IHNV病毒核酸的最低检出量为100pg,且特异性高,与其他病毒无交叉反应。应用建立的液相芯片技术检测鱼类中IHNV,并与RT-PCR方法进行比较,检测结果基本一致,但该方法比RT-PCR法更灵敏。表明初步建立了检测IHNV的液相芯片技术,为进一步构建其他水生动物病原体快速高通量检测平台提供参考。

A liquid chip assay was developed to detect infectious haematopoietic necrosis virus（IHNV）. The IHNV N gene in the Gen Bank was analyzed using the software DNAStar 7.0. Specific IHNV primers labeled with biotin was prepared and coupled with fluorescence-coded microspheres. The amino modified probe was used for hybridization with PCR products of IHNV,then fluorescence signal in the reaction system was detected using BD FACS Array. The results showed that IHNV could be accurately detected by the developed liquid chip assay. Its detection limit was 100 pg viral RNA and its specificity was very high without cross reaction with other viruses.The IHNV liquid chip assay was used to detect IHNV in fish samples and compared with the conventional RT-PCR resulting in basically the same,but more sensitive than RT-PCR. The development of IHNV liquid chip assay provided references for further research on rapidly high throughput detection for other aquatic pathogens with the technique。