| 马媾疫锥虫SYBR GreenⅠ荧光定量PCR检测方法的建立 |
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| 引用本文： | 王群,郑小龙,朱来华,肖西志,邓明俊,岳志芹,孙涛,赵玉然.马媾疫锥虫SYBR GreenⅠ荧光定量PCR检测方法的建立[J].中国预防兽医学报,2012,34(9):724-727. |
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| 作者姓名： | 王群 郑小龙 朱来华 肖西志 邓明俊 岳志芹 孙涛 赵玉然 |
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| 作者单位： | 山东出入境检验检疫局,山东青岛,266002 |
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| 摘 要： | 为建立一种特异、快速的媾疫锥虫检测方法,本研究通过PCR方法从马媾疫锥虫动基体基因组中扩增得到395 bp的特异的保守序列,将其克隆到pMD-18T载体中构建重组质粒标准品,以10倍倍比稀释的质粒标准品为模板,进行SYBR GreenⅠ荧光定量PCR扩增并制作标准曲线,建立马媾疫锥虫荧光定量PCR的检测方法。结果表明:该方法的检测灵敏度可达到1拷贝/μL,并且与马属动物其他传染病无交叉反应。其组间及组内变异系数分别小于3.183%和3.842%。该方法的建立为快速及特异性检测马媾疫锥虫提供了有效的方法。
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| 关 键 词： | 马媾疫锥虫 SYBR GreenⅠ 荧光定量 |
Development of SYBR Green Ⅰ fluorescent quantitation assay for detection of Trypanosoma equiperdum |
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| WANG Qun
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ZHENG Xiao-long
,
ZHU Lai-hua
,
XIAO Xi-zhi
,
DENG Ming-jun
,
YUE Zhi-qin
,
SUN Tao
,
ZHAO Yu-ran.Development of SYBR Green Ⅰ fluorescent quantitation assay for detection of Trypanosoma equiperdum[J].Chinese Journal of Preventive Veterinary Medicine,2012,34(9):724-727. |
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| Authors: | WANG Qun ZHENG Xiao-long ZHU Lai-hua XIAO Xi-zhi DENG Ming-jun YUE Zhi-qin SUN Tao ZHAO Yu-ran |
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| Institution: | (Shandong Entry-Exit Inspection and Quarantine Bureau,Qingdao 266002,China) |
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| Abstract: | To develop a SYBR GreenⅠreal-time PCR assay(qPCR) to detect Trypanosoma equiperdum,a 395 bp specific and consensus DNA sequence was amplified from the kinetoplast of T.equiperdum and cloned into plasmid pMD-18T.Standard curve was generated by using serial dilutions of recombinant plasmid.Detection results indicated that the assay has high sensitivity with a detection limit of 1 copies per reaction and no cross-reaction with other equine pathogens.Those results indicated that a convenient,specific,high performance and sensitive qPCR assay was established successfully for the detection of T.equiperdum. |
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| Keywords: | Trypanosoma equiperdum SYBR GreenⅠ fluorescent quantitation |
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