鸭圆环病毒实时荧光定量PCR检测方法的建立及初步应用
发布时间：2018-08-19 16:58
【摘要】：为实现对鸭圆环病毒(DuCV)的快速检测,分析比对Gen Bank中登录的DuCV全基因组序列,在其保守区设计并合成一对特异性引物,将扩增的212 bp片段克隆入pMD-18T载体中,获得pMD18DuCV重组质粒,通过SYBR GreenⅠ荧光掺入法分别对系列稀释pMD18DuCV中DuCV特异性片段进行扩增,通过pMD18DuCV浓度对数与Ct值制作了标准曲线并推导出直线回归方程,二者相关系数(R~2)为0.9968,建立了DuCV实时荧光定量PCR检测方法,重复性和灵敏性检测结果表明重复性良好,检测下限为67.2拷贝/μL;将建立的DuCV实时荧光定量PCR检测方法分别对鸭肝炎病毒(DHV)、大肠杆菌、新城疫病毒(NDV)、H9亚型禽流感病毒、小鹅瘟病毒(GPV)核酸或cDNA进行特异性检测,结果表明特异性良好。初步应用结果表明,建立的DuCV实时荧光定量PCR方法检出率为26.4%(62/235),明显优于常规PCR检测结果15.3%(36/235)。
[Abstract]:In order to detect duck circovirus (DuCV) quickly, a pair of specific primers were designed and synthesized in the conserved region of Gen Bank to analyze and compare the whole DuCV genome sequence registered in Gen Bank. The amplified 212bp fragment was cloned into pMD-18T vector to obtain pMD18DuCV recombinant plasmid. The specific fragments of DuCV in diluted pMD18DuCV were amplified by SYBR Green 鈪，
本文编号：2192267
[Abstract]:In order to detect duck circovirus (DuCV) quickly, a pair of specific primers were designed and synthesized in the conserved region of Gen Bank to analyze and compare the whole DuCV genome sequence registered in Gen Bank. The amplified 212bp fragment was cloned into pMD-18T vector to obtain pMD18DuCV recombinant plasmid. The specific fragments of DuCV in diluted pMD18DuCV were amplified by SYBR Green 鈪，
本文编号：2192267
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