摘要
目的建立2种实时荧光定量PCR方法,用于在血液样本中检测有可能威胁血液安全的人巨细胞病毒(human cytomegalovirus,HCMV)。方法根据Gen Bank中HCMV的MIE基因保守序列的分析结果,分别建立基于SYBR Green和Taq Man探针的2种实时定量PCR检测方法,同时用这2种方法对来自血液制品厂的血浆进行检测。结果基于SYBR Green的实时定量PCR检测方法,得到的熔解曲线峰单一,建立的2种实时定量PCR方法对于HCMV的DNA具有高度特异性和灵敏度,检测下限均为50 copies/m L,并且具有较好的批间和批内重复性。结论应用本研究建立的实时荧光定量PCR方法对献浆人群血浆样品进行HCMV检测,2种方法检测结果一致,证明该方法可用于在血液样本中检测HCMV。
Objective To establish two quantitative real-time PCR methods in detecting Human Cytomegalovirus （HCMV） which would be a threat to blood safety. Methods According to the conserved gene MIE of HCMV available in Gen-Bank, two quantitative real-time PCR methods based on SYBR Green and TaqMan probes were established for the detection of HCMV. The plasma from blood products provided by manufacturer were detected using the established real-time Q-PCR assays. Results A single melting curve indicated a single predominant product in SYBR Green real-time Q-PCR method. The specificity and sensitivity of the established real-time Q-PCR assays were high and there were no cross reactivity with HBV and syphilis. The detection limit for both quantitative real-time PCR methods was 50 copies/ml. In the duplicated ex- periment, the replication of intra-assay and inter-assay was good. Conclusion The real-time Q-PCR methods with good specificity, sensitivity and consistency could be used for detecting HCMV infection and viral loads in plasma.
出处
《中国输血杂志》
CAS
北大核心
2015年第6期637-641,共5页
Chinese Journal of Blood Transfusion
