国际间实验室禽流感能力验证样品制备技术的研究与应用

Preparation and Testing of International Proficiency Testing Samples for Detection of Avian Influenza

通过对禽流感病毒基因序列的比较分析,在其血凝素(HA)基因保守区,设计引物,通过基因克隆,体外转录等技术,制备出病毒HA基因的RNA片段,并对体外转录的RNA进行了定量、稀释,以作为用于国际间实验室禽流感能力验证的样品,并建立了绝对定量实时荧光RT-PCR检测方法。样品的均匀性和稳定性实验结果证明了样品制备技术的可行性,并为我国首次成功组织实施国际间实验室禽流感能力验证计划奠定了基础。

According to the gene comparison and analysis of H5N1 avian influenza virus, primers were designed targeting hemagglutinin （HA） gene of the virus. And the RNA fragments, obtained from gene cloning and in vitro transcription, were quantitated and diluted as international proficiency testing samples. And an absolute quantitative real-time RT-PCR assay （AQ-RT-PCR） was also developed. Then the homogeneity testing and stability testing were performed. Results of homogeneity testing and stability testing showed that the samples suits the proficiency testing, and the technique of samples preparation is feasible, which lay the foundation for our organization and fulfillment of the international proficiency testing for detection of Avian Influenza for the first time.