[1]林裕胜,江锦秀,江 斌,等.肺炎亚种多重PCR检测方法的建立[J].中国预防兽医学报,2018,(09):812-817.[doi:0.3969/j.issn.1008-0589.201710026]
 LIN Yu-sheng,JIANG Jin-xiu,JIANG Bin,et al.Development of a triplex PCR assay for detection of Mycoplasma ovipneumoniae, M.mycoides subsp. Capri and M.capricolum subsp. capripneumoniae[J].Chinese journal of preventive veterinary medicine,2018,(09):812-817.[doi:0.3969/j.issn.1008-0589.201710026]
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肺炎亚种多重PCR检测方法的建立()
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《中国预防兽医学报》[ISSN:1008-0589/CN:23-1417/S]

卷:
期数:
2018年09
页码:
812-817
栏目:
诊断和检测技术
出版日期:
2018-09-17

文章信息/Info

Title:
Development of a triplex PCR assay for detection of Mycoplasma ovipneumoniae, M.mycoides subsp. Capri and M.capricolum subsp. capripneumoniae
文章编号:
1008-0589(2018)9-0812-06
作者:
 

林裕胜江锦秀江 斌游 伟胡奇林*

 (福建省农业科学院 畜牧兽医研究所,福建 福州 35001
Author(s):
 

LIN Yu-sheng JIANG Jin-xiu JIANG Bin YOU Wei HU Qi-lin*

 (Institute of Animal Husbandry & Veterinary Medicine, Fujian Academy of Agricultural Sciences, Fuzhou 350013, China)
关键词:
绵羊肺炎支原体丝状支原体山羊亚种山羊支原体山羊肺炎亚种多重PCR
分类号:
S852.61
DOI:
0.3969/j.issn.1008-0589.201710026
文献标志码:
A
摘要:
为同时快速检测绵羊肺炎支原体(Mo)、丝状支原体山羊亚种(Mmc)和山羊支原体山羊肺炎亚种(Mccp),本研究根据Mo的P80基因,Mmc的MLC_1760和Mccp的arcA基因序列,设计合成了3对特异性引物,建立了同时检测Mo、Mmc和Mccp的多重PCR方法。该方法可以同时扩增出Mo、Mmc和Mccp的特异性片段,而对大肠杆菌、巴氏杆菌、羊传染性脓疱病毒等常见羊病病原无交叉反应,对Mo、Mmc和Mccp的最低检出量分别为3.62×104拷贝/μL、4.03×105拷贝/μL和6.78×105拷贝/μL。应用该方法对75份临床样品进行检测,结果与单一PCR检测结果一致。对32份临床样品同时用该多重PCR方法与分离鉴定法进行检测,结果显示,两种方法对Mo、Mmc、Mccp的检测符合率分别为87.5 %、100 %、100 %。本研究建立的多重PCR方法特异性强、敏感性较高,适用于Mo、Mmc和Mccp临床快速检测及流行病学调查。
  
Abstract:
In order to simultaneously and rapidly detect Mycoplasma ovipneumoniae (Mo), Mycoplasma mycoides subsp. capri (Mmc) and Mycoplasma capricolum subsp. capripneumoniae (Mccp), a triplex PCR assay was developed with three pairs of specific primers designed according to the P80 gene of Mo, MLC_1760 gene of Mmc and arcA gene of Mccp. The specific PCR products of Mo (705bp), Mmc (317bp) and Mccp (242bp) were simultaneously amplified from the DNAs of Mo, Mmc and Mccp, respectively, by the triplex PCR, but had no cross reaction with E.coli, Pm, ORFV and other goats and sheep pathogens. The detection limits for Mo, Mmc and Mccp were 3.62×104copies/μL, 4.03×105copies/μL and 6.78×105copies/μL, respectively. Using the assay to detect 75 clinical samples collected from Fujian, the results showed that the triplex PCR was 100% agreement with the single PCR. The coincidence rates between the triplex PCR assay and Mycoplasma isolation were 87.53% for Mo, 100% for Mmc, 100% for Mccp, respectively, from 30 clinical samples. These results indicated that the triplex PCR assay is specific and sensitive, and is suitable for the routine detection of the Mo, Mmc, Mccp and their epidemiological investigation.

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                                        (本文编辑:李   娜)

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更新日期/Last Update: 2018-09-28