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Position: Home > Articles > Development and Application of Nested PCR Assay for Detection of Dairy Cattle-Derived Cyclospora sp. Scientia Agricultura Sinica 2007,40 (9) 2073-2078

牛源环孢子虫套式PCR检测方法的建立及初步应用

作  者：
肖淑敏;李国清;李韦华;周荣琼;杨建伟
单  位：
华南农业大学兽医学院
关键词：
环孢子虫;奶牛;套式PCR;检测
摘  要：
【目的】建立牛源环孢子虫套式PCR检测方法并初步应用于临床检测。【方法】根据牛源环孢子虫18S rDNA序列,设计1对保守引物和1对特异性引物,建立牛源环孢子虫套式PCR检测方法。通过阳性参照摸索出该检测方法的最佳反应条件,进行特异性和敏感性试验。并对168份临床样品进行了检测。【结果】该套式PCR能特异性地扩增出牛源环孢子虫基因组DNA中相应片段,与艾美耳球虫、隐孢子虫、贾第虫等10种相关原虫基因组DNA无交叉反应;最低能检测到2.85×10-2fg的阳性参照DNA;对临床样品检测结果表明PCR技术检查阳性者显微镜检查都为阳性。【结论】建立的套式PCR方法具有高度的特异性和敏感性,有较好的临床应用价值。
译  名：
Development and Application of Nested PCR Assay for Detection of Dairy Cattle-Derived Cyclospora sp.
作  者：
XIAO Shu-min1,2,LI Guo-qing1,LI Wei-hua1,ZHOU Rong-qiong1,YANG Jian-wei1(1College of Veterinary Medicine,South China Agricultural University,Guangzhou 510642;2School of Safety Science and Engineering,Henan Polytechnic University,Jiaozuo 454000)
关键词：
Cyclospora sp.;Dairy cattle;Nested PCR;Detection
摘  要：
【Objective】 To develop a nested PCR assay for detection of cattle-derived Cyclospora sp.【Method】 Based on the cattle-derived Cyclospora sp.sequences,two pairs of primers were designed.These primers selectively amplified a 168-bp DNA fragment of the 18S rRNA gene of cattle-derived Cyclospora sp.With these primers,a nested PCR assay for detection of cattle-derived Cyclospora sp.was developed.【Result】The nested PCR assay was specific and there is no cross-reaction with other parasites,such as Eimeria spp.,Cryptosporidium spp.,Giardia sp.,Toxoplasma sp.,Trichuris sp.and cattle ciliate.The assay was able to detect as low as 2.85×10-2 fg of control positive DNA.The results of detection of clinical samples revealed that the assay was coinciding with microscopic examination.【Conclusion】The nested PCR assay was specific and sensitive and it will be an effective tool for detection of Cyclospora sp.in cattle feces.

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