| 牛支原体、无乳支原体和丝状支原体丝状亚种小克隆三重PCR检测方法的建立及初步应用 |
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| 引用本文： | 李大伟,黄灿平,张彦明,谢建华,冉智光,熊仲良,范伟兴.牛支原体、无乳支原体和丝状支原体丝状亚种小克隆三重PCR检测方法的建立及初步应用[J].畜牧兽医学报,2011,42(2). |
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| 作者姓名： | 李大伟 黄灿平 张彦明 谢建华 冉智光 熊仲良 范伟兴 |
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| 作者单位： | 1. 中国动物卫生与流行病学中心,青岛,266032
2. 重庆市动物疫病预防控制中心,重庆,401147 |
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| 基金项目： | 科技部863专项基金项目 |
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| 摘 要： | 本研究旨在建立一种可一次性区分牛支原体、丝状支原体丝状亚种小克隆和无乳支原体的三重PCR诊断方法,为临床诊断和流行病学调查提供可靠检测技术.根据GenBank发表的上述3种病原的基因组序列保守区域设计3对特异性引物建立三重PCR方法;确定其检测敏感性,以猪支原体、鸡支原体、无乳支原体和丝状支原体丝状亚种小克隆基因作模板检验其特异性;同时和病原分离鉴定结果对比其准确性.结果表明在优化体系和条件下能够同时得到扩增长度为448、549、375 bp 3条特异性片段,未扩增出猪、鸡支原体模板特异性片段;其敏感性(可检测到的最小模板DNA含量)为0.8 ng·μL-1;36份临床样品检测结果显示,三重PCR检测结果与分离培养鉴定方法一致,均能鉴定出牛支原体阳性病料.本研究建立的三重PCR诊断方法能够一次性鉴别3种支原体,具有高敏感性、特异性和准确性,可用于临床诊断和流行病学调查.
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| 关 键 词： | 牛支原体 丝状支原体丝状亚种小克隆 无乳支原体 三重PCR |
Establishment and Application of the Triplex PCR for Detecting M.Bovis, M.mycoides subsp.mycoides Small Colony Type and M.agalactiae |
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| LI Da-wei,HUANG Can-ping,ZHANG Yan-ming,XIE Jian-hua,RAN Zhi-guang,XIONG Zhong-liang,FAN Wei-xing.Establishment and Application of the Triplex PCR for Detecting M.Bovis, M.mycoides subsp.mycoides Small Colony Type and M.agalactiae[J].Acta Veterinaria et Zootechnica Sinica,2011,42(2). |
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| Authors: | LI Da-wei HUANG Can-ping ZHANG Yan-ming XIE Jian-hua RAN Zhi-guang XIONG Zhong-liang FAN Wei-xing |
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| Institution: | LI Da-wei1,HUANG Can-ping1,ZHANG Yan-ming1,XIE Jian-hua2,RAN Zhi-guang2,XIONG Zhong-liang2,FAN Wei-xing1 (1.China Animal Health and Epidemiology Center,Qingdao 266032 China,2.ChongqingCenter for Animal Disease Control and Prevention,Chongqing 401147,China) |
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| Abstract: | The study was aimed at establishing a triplex PCR to differentiate M.bovis,M.mycoides subsp.mycoides Small Colony Type and M.agalactiae out of the same sample,for clinical diagnosis and epidemiological survey.Three pairs of primers were designed and synthesized according to the published literatures in GenBank.And a triple PCR was developed to amplify conservative regions of each bacterial genomes of M.Bovis,M.mycoides subsp.mycoides Small Colony Type and M.agalactiae,with the target sequences of 448,549 an... |
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| Keywords: | M bovis M mycoides subsp mycoides Small Colony Type M agalactiae triplex PCR |
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