摘要
目的建立快速、灵敏、特异及检测结果易判断的PCR方法,并应用于大规模猕猴种群的弓形虫常规检测中。同时比较巢式PCR和单一PCR的一致性。方法根据弓形虫保守基因p30(SAG1)设计了内、外两对进行巢式PCR扩增以及B1基因设计一对引物进行单一PCR扩增,将DNA样本进行10倍倍比稀释,以检测巢式PCR反应的灵敏度;并对医学生物学研究所自繁猕猴共150只进行了弓形虫检测。结果巢式PCR检测法检测限度可达10-3ng/uL,而且方法特异。两种PCR法检测结果基本一致,其中巢式PCR检测阳性率(10%)稍高于单一PCR检测阳性率(8.67%)。结论巢式PCR和一次PCR方法都可应用于猕猴弓形虫的常规检测中,并提示巢式PCR比单一PCR更敏感、检出率更高。
Objective To establish a rapid, sensitive, specific PCR technique for detection of toxoplasma gondii and applying in breeding rhesus macaques. The results from nested-PCR and single-step PCR assay were compared. Methods Two pairs of primers matched with P30 gene were designed for nested PCR and B1 gene primers were amplified in single-step PCR. Toxoplasma gondii DNA samples were serially tenfold diluted to determine the sensitivity of the nested PCR. 150 samples of breeding rhesus macaques from the Institute of Medical Biology were assayed by those two PCR methods. Results The nested PCR was a specific method and its sensitivity was 10-3 ng/μL. The results of nested PCR were almost consistent with that of single-step PCR assay, but the positive rate （10 % ） was higher than that （8.67 % ） of single-step PCR. Conclusions Both the two PCR methods can be used to detect toxoplasma gondii DNA from monkey samples, and the nested PCR technique is more sensitive than single-step PCR.
出处
《中国比较医学杂志》
CAS
2008年第6期20-24,共5页
Chinese Journal of Comparative Medicine
