中国寄生虫学与寄生虫病杂志 ›› 1991, Vol. 9 ›› Issue (2): 90-93.

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质粒PF rep20探针检测我国恶性疟原虫的初步研究

刁忠玉,陈培霞,李霖,吴灿理,陈南春   

  1. 第四军医大学寄生虫学教研室; 第四军医大学寄生虫学教研室; 第四军医大学寄生虫学教研室; 第四军医大学寄生虫学教研室; 第四军医大学生物化学教研室
  • 收稿日期:2017-01-09 修回日期:2017-01-09 出版日期:1991-05-31 发布日期:2017-01-09

DETECTION OF PLASMODIVM FALCIPARUM USING THE PLASMID PF REP 2O PROBE (PRELIMINARY REPORT)

  • Received:2017-01-09 Revised:2017-01-09 Online:1991-05-31 Published:2017-01-09

摘要: 用~(33)P中标记的质粒PF rep20探针以DNA-DNA斑点杂交检测5种疟原虫,仅恶性疟原虫P.f.呈现阳性反应,间日疟原虫、约氏疟原虫、食蟹猴疟原虫及诺氏疟原虫不发生杂交;~(32)P标记的P.f.海南株基因组DNA探针与上述诸疟原虫均可发生杂交。基因诊断海南省和云南省恶性疟患者血样本39份的杂交阳性率,质粒探针和基因探针分别为94.9%和97.4%。检测人工培养的海南株、云南株和安徽株P.f.敏感度,质粒探针均为0.001%原虫血症和10Pg原虫DNA,基因组探针对海南株原虫为0.0001%和1Pg。提示质粒PF rep20探针种的特异性强,敏感性较高,似可用于我国恶性疟原虫检测。

关键词: 恶性疟原虫, 脱氧核糖核酸, 基因诊断

Abstract: A cloned repetitive DNA sequence (rep 20) labeled with 32P was evaluated as diagnostic probe for P. falciparum in 39 blood samples from Hainan and Yunan provinces. Its specificity and sensitivity were studied and compared with total genomic DNA probe isolated from P. f. Hainan Fccj/HN isolate. The pPF rep 20 probe recognized the DNA of P. f. Hainan, Yunnan and Anhui isolates, but did not hybridize with the DNA of P. vivax, P. cynomolgl, P. knowlesi and P. yoelii. The genomic probe hybridized with the DNA of 3 isolates and cross-hybridized with the DNA of all the other Plasmodium species tested, but did not hybridize with host DNA. The plasmid rep 20 probe was able to detect para-sitemia level of 0.001% in 20μl blood from culture and 10pg DNA of 3 P. f. isolates after 3 days film exposure. It could hybridize with the blood samples of P. f. patients from Hainan and Yunnan with a sensitivity of 95% (37/39). The genomic probe could detect the same parasitemia and DNA levels as plasmid rep 20 probe for Yunnan and Anhui isolates, and 0.0001% parasitemia and 1 pg DNA for Hainan isolate. It had a sensitivity of 97% (38/39) when used to detect patient samples. The results indicated that plasmid rep 20 probe was specific and rather sensitive to DNA of P. falciparum isolates from China and may be useful in epidemiological studies.