羊附红细胞体单管套式PCR检测方法的建立及初步应用  

Establishment and Preliminary Application of a Single-tube Nested PCR Assay for Detection of Goat Eperythrozoon

作　　者：李丽[1] 王书全[1] 

机构地区：[1]辽宁医学院畜牧兽医学院,辽宁锦州121001

出　　处：《动物医学进展》2013年第5期87-90,共4页Progress In Veterinary Medicine

摘　　要：为了建立绒山羊附红细胞体单管套式PCR检测方法,根据GenBank上发表的羊附红细胞体16S rRNA基因序列,设计内、外2对特异性引物,建立单管套式PCR方法,同时进行了特异性、敏感性试验及临床检测。结果表明,本检测方法能够扩增基因片段大小为681bp,与GenBank收录的相关序列同源性为97.6%～99.2%,而与牛温氏附红细胞体、猪附红细胞体、绵羊肺炎支原体、猪肺炎支原体、大肠埃希菌相关病原基因组均无交叉反应,能够检测出DNA的最小量为14.8fg。对74份临床样品进行了检测,阳性率为43.2%,高于常规PCR和血涂片镜检测结果,表明本检测方法具有较高的敏感性,可用于羊附红细胞体感染的早期诊断,有较高的临床应用价值。To establish a single-tube nested PCR assay for detection to Liaoning Cashmere goat Eperythrozoon, according to the published Eperythrozoon 16 S rRNA gene sequence in GenBank, two pairs of primers were designed to establish nested PCR assay for detecting the disease caused by goat Eperythrozoon. The results of verification experiments showed that the targeted gene fragment was 681 bp in length and homol- ogy of nested PCR product was 97.6 ^-99.2 ~, and this detection method had no cross reaction with M. wenyoni, Eperythrozoon suis, Sheep Mycoplasrna pneumonia, Swine Mycoplasrna pneumonia, E. coll. The sensitivity reached to 14.8 fg DNA. The 43.2~//oo positive rate in 74 clinical samples by the single-tube nes- ted PCR assay was higher than those by routine PCR and blood smear. The results revealed that the single- tube nested PCR assay had higher specificity, sensitivity, and can be used as an effective detection method of goat Eperythrozoon.

关 键 词：羊附红细胞体 单管套式PCR 诊断技术 

分 类 号：S853.32[农业科学—临床兽医学]