鸡减蛋综合征病毒Hexon基因SYBR GreenⅠ实时荧光定量PCR检测方法的建立  

Detection of Hexon gene from egg drop syndrome virus using SYBR GreenⅠquantitative real time PCR technology

作　　者：杜冬华 周静[1] 王爱华[1] 孙继国[2] 

机构地区：[1]河北北方学院动物科技学院,河北张家口075131 [2]河北农业大学动物科技学院,河北保定071000

出　　处：《中国兽医学报》2014年第8期1231-1234,共4页Chinese Journal of Veterinary Science

基　　金：河北张家口市科学技术和地震局计划资助项目(12110037C-4)

摘　　要：用PCR方法扩增出鸡减蛋综合征病毒(EDSV)Hexon基因保守片段,经琼脂糖凝胶电泳及序列测定分析扩增产物的特异性。以构建的阳性重组质粒作为标准品,建立SYBR GreenⅠ实时荧光定量PCR反应的扩增曲线和溶解曲线,并绘制标准曲线。结果表明,建立的EDSV荧光定量PCR标准曲线Ct值与1×101～1×106拷贝/μL的基因拷贝数呈现良好线性关系,灵敏度可达10拷贝,且特异性及重复性良好;说明本试验建立的SYBR GreenⅠ实时荧光定量PCR检测方法可用于EDSV的诊断及病原的定量分析。The conservative fragment of Hexon gene from egg drop syndrome virus（EDSV） was amplified by PCR and analysis of the amplification specificity by agarose gel electrophoresis and sequencing method. The recombinant plasmid was used to establish melting and standard curves of the SYBR Green I quantitative real time PCR. The standard curve for the viral genomic copy number and threshold cycle ranging from 1×10^1～1×10^6 copies/μL were linear. Sensitivity of the detection method was 10 copies,and have a good specificity and repeatability. These data indicated that the SYBR Green I quantitative real time PCR can be used for EDSV diagnostics and quantification.

关 键 词：鸡减蛋综合征病毒 荧光定量PCR SYBR GreenⅠ 

分 类 号：S852.65[农业科学—基础兽医学]