牛源贾第虫套式PCR检测方法的建立

目的建立牛源贾第虫套式PCR检测方法。方法根据牛源贾第虫TPI基因序列,设计1对保守引物和1对特异性引物,建立套式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;并用临床样品进行验证。结果该套式PCR能特异性地扩增出牛源贾第虫基因组DNA中相应片段,与艾美耳球虫、隐孢子虫、环孢子虫、弓形虫、毛首线虫和纤毛虫等10种相关原虫基因组DNA无交叉反应;最低能检测到0.395fg的阳性参照DNA。结论建立的套式PCR方法具有高度的特异性和敏感性,可以用于临床样品的检测。

Development of the nested PCR assay for detection of dairy cattle-derived Giardia lamblia

To develop a nested PCR assay for detection of cattle-derived Giardia lamblia, two pairs of primers were designed based on the cattle-derived G. lamblia sequences. These primers selectively amplified a 357-bp DNA fragment of the triose phosphate isomerase (TPI) gene of cattle-derived G. lamblia. With these primers, a nested PCR assay for detection of cattle-derived G. lamblia was developed. The results showed that the nested PCR assay was specific and there was no cross-reaction with other parasites, such as Eimeria spp., Cryptosporidium spp., Cyclospora sp., Toxoplasma gondii, Trichuris sp. and cattle ciliate. The assay was able to detect as low as 0.395 fg of positive control DNA. The results of detection of clinical sample revealed that the assay was veracious and it will be an effective tool for detection of G. lamblia in cattle feces.