基孔肯雅病毒荧光定量PCR检测方法的建立

Real-time PCR method for rapid detection of Chikungunya virus

目的建立一种快速、敏感、特异的实时荧光定量PCR方法,检测基孔肯雅病毒。方法通过序列比对挑选出基孔肯雅病毒基因组中高度保守的序列,在此序列上设计引物及TaqMan探针,建立实时荧光定量PCR反应体系。结果经优化的荧光定量PCR方法有较好的灵敏度和特异性,对阳性对照质粒标准品的灵敏度可达21拷贝/μl,通过检测与传播媒介相似的流行性乙型脑炎病毒、黄热病毒、登革热病毒无交叉反应。结论该方法的建立在基孔肯雅热的疾病防控方面有较好的应用前景。

Objective To establish a rapid,sensitive and specific detection method for detecting Chikungunya virus（CHIKV） using Real-time fluorescence quantitative PCR.Methods With specifically designed primers and a TaqMan probe on the highly conserved sequence of CHIKV through alignment,the sensitivity of the Real-time fluorescence quantitative PCR assay was optimized by evaluating different concentrations of primers and probes.Results A specific Real-time PCR method was developed with the sensitivity of 21 copies/μl for CHIKV,a synthetic CHIKV genome DNA as a positive control;Japanese encephalitis virus,Yellow fever virus,Dengue virus were using to examine the specificity.Conclusion Promising prospects of this assay could be expected for Chikungunya fever prevention and control.