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使用TaqMan®实时荧光定量PCR技术快速检测伤寒沙门氏菌。

Use of TaqMan® real-time PCR for rapid detection of Salmonella enterica serovar Typhi.

作者信息

Ranjbar Reza, Naghoni Ali, Farshad Shohreh, Lashini Hadi, Najafi Ali, Sadeghifard Nourkhoda, Mammina Caterina

机构信息

Baqiyatallah University of Medical Sciences Molecular Biology Research Center Tehran Iran.

Shiraz University of Medical Sciences Prof. Alborzi Clinical Microbiology Research Center Shiraz Iran.

出版信息

Acta Microbiol Immunol Hung. 2014 Jun;61(2):121-30. doi: 10.1556/AMicr.61.2014.2.3.

Abstract

We evaluated the performances of a newly designed real-time polymerase chain reaction (PCR) assay using TaqMan® probes to detect Salmonella Typhi. TaqMan® real-time PCR assays were performed by designed primers and probe based on the staG gene for detecting S. Typhi. The specificity of the assay was evaluated on 15 Salmonella serovars. The analytical specificity was evaluated on 20 non-Salmonella microorganisms. The analytical sensitivity was assessed using decreasing DNA quantities of S. Typhi ATCC 19430. Finally the detection capability of the TaqMan® real-time PCR assay on isolates recovered from patients with Salmonella infections was compared to the conventional PCR assay. Only S. Typhi strain had positive results when subjected to the assay using Typhi-specific real-time PCR. No amplification products were observed in real-time PCR with any of the non-Salmonella microorganisms tested. The TaqMan® real-time PCR was more sensitive than the conventional PCR. In conclusion, we found that the easy-to-use real-time PCR assays were faster than conventional PCR systems. The staG-based TaqMan® real-time PCR assay showed to be specific and sensitive method for the safe and rapid detection of the S. Typhi.

摘要

我们评估了一种新设计的使用TaqMan®探针的实时聚合酶链反应(PCR)检测伤寒沙门氏菌的性能。基于staG基因设计引物和探针,进行TaqMan®实时PCR检测以检测伤寒沙门氏菌。在15种沙门氏菌血清型上评估了该检测方法的特异性。在20种非沙门氏菌微生物上评估了分析特异性。使用逐渐减少的伤寒沙门氏菌ATCC 19430 DNA量评估了分析灵敏度。最后,将TaqMan®实时PCR检测方法对从沙门氏菌感染患者中分离出的菌株的检测能力与传统PCR检测方法进行了比较。使用伤寒沙门氏菌特异性实时PCR检测时,只有伤寒沙门氏菌菌株得到阳性结果。在对任何测试的非沙门氏菌微生物进行的实时PCR中均未观察到扩增产物。TaqMan®实时PCR比传统PCR更灵敏。总之,我们发现这种易于使用的实时PCR检测方法比传统PCR系统更快。基于staG的TaqMan®实时PCR检测方法是一种安全、快速检测伤寒沙门氏菌的特异性和灵敏方法。

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