DEVELOPMENT OF A DIAGNOSTIC-TEST FOR YERSINIA-PESTIS BY THE POLYMERASE CHAIN-REACTION
DEVELOPMENT OF A DIAGNOSTIC-TEST FOR YERSINIA-PESTIS BY THE POLYMERASE CHAIN-REACTION
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DOI：
10.1111/j.1365-2672.1994.tb01622.x
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发表时间：
1994-03-01
期刊：
影响因子：
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通讯作者：
DROSDOV, IG
中科院分区：
文献类型：
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作者：
NORKINA, OV;KULICHENKO, AN;DROSDOV, IG
以100kb(Pfra)和10kb(PPst)染色体外复制子携带的501bpCAF1基因片段和443bpplA基因片段为靶标，研究了聚合酶链式反应(PCR)扩增技术检测和鉴定纯培养物和生物样品细胞裂解物中鼠疫菌DNA的条件。用鼠疫EV粗细胞裂解物进行聚合酶链式反应的灵敏度为10-50CFU。将鼠疫EV靶细胞与含0.4%柠檬酸钾的小白鼠新鲜血液混合，根据样品制备方法的不同，可检测到400~100CFU ml(-1)的血液。在我们的试验中，聚合酶链式反应能有效地检测实验感染鼠疫杆菌KM638毒株的小白鼠血液中的耶尔森氏菌。该方法可用于鼠疫耶尔森氏菌的常规检测和鉴定。
A 501 bp caf1 gene fragment and a 443 bp of pla gene fragment carried by 100 kb (pFra) and 10 kb (pPst) species-specific extrachromosomal replicons, respectively, were used as targets to study the conditions under which DNA amplification by polymerase chain reaction (PCR) may be applied to detect and identify Yersinia pestis DNA in cell lysates of pure cultures and biological samples. The sensitivity limit of PCR with the crude cell lysates of Y. pestis EV was estimated as 10-50 cfu in reaction mixture. When target Y. pestis EV cells were mixed with fresh blood of white mice, which contained 0.4% potassium citrate, the PCR detection level varied from 400 to 100 cfu ml(-1) of blood depending on the method used for preparing the sample. In our tests PCR was effective for the detection of yersinia in the blood of white laboratory mice experimentally infected with virulent Y. pestis KM638 strain. This method can be considered convenient for routine detection and identification of Y. pestis.