摘要
目的建立基于TpN17基因检测血液中梅毒螺旋体的荧光定量PCR方法。方法根据梅毒螺旋体TpN17基因保守区域设计引物,构建质粒标准品pTpN17,优化定量PCR反应体系检测该体系的灵敏度和特异性;同时,采用该方法对梅毒确证阳性的血液样本进行检测。结果研究表明,基于TpN17基因的SYBR GREEN荧光定量PCR方法可以有效检测血液中10^6-10^1 copies/μL的梅毒螺旋体并通过熔解曲线判定结果的特异性。结论以TpN17基因为扩增目的基因的SYBR GREEN荧光定量PCR方法具有灵敏快速便宜的特点,适用于在血液检测梅毒螺旋体,能广泛应用于献血及血液样本检测等领域,提高我国的血液安全。
Objective To establish a real-time PCR method to detect Treponema pallidum in blood samples based on the detection of TpN17 gene. Methods The primers were designed according to the conserved domain of TpN17 gene and the recombinant vector named pTpN17 was used as the standard template. The real-time PCR reaction system was optimized focusing on specificity and sensitivity. Afterwards ,the assay based on this method for quick detection of confirmed T. paUidum in serum from blood donors was performed to evaluate the validity of this real-time PCR system. Results SYBR GREEN fluo- rescent were positive for all detections of TpN17 gene in the specimens collected from T. pallidum infected blood donors. The range of detections is from 10^6 - 10^1 copies/μl identified through melting curve analysis. Conclusion The real-time PCR system for detecting T. pallidum has been established with the characteristics of low price, sensitivity and specificity. This method could be applied in many T. paUidum detection fields such as blood screening and blood specimen detection, by implementing which would benifit the blood safety of our country.
出处
《中国输血杂志》
CAS
CSCD
北大核心
2014年第7期722-724,共3页
Chinese Journal of Blood Transfusion
