development and inter-laboratory validation of unlabeled probe melting curve analysis for detection of jak2 v617f mutation in polycythemia vera开发和实验室内部验证标记探针融化曲线分析检测jak2 v617f突变真性红细胞增多.pdfVIP

Development and Inter-Laboratory Validation of Unlabeled Probe Melting Curve Analysis for Detection of JAK2 V617F Mutation in Polycythemia Vera 1. 2. 3 1 4 5 1,3,4 Zhiyuan Wu , Hong Yuan , Xinju Zhang , Weiwei Liu , Jinhua Xu , Wei Zhang , Ming Guan * 1 Department of Laboratory Medicine, Huashan Hospital, Shanghai Medical College, Fudan University, Shanghai, People’s Republic of China, 2 Department of Clinical Laboratory, The First Affiliated Hospital of Dalian Medical University, Dalian, People’s Republic of China, 3 Central Laboratory, Huashan Hospital, Shanghai Medical College, Fudan University, Shanghai, People’s Republic of China, 4 Department of Dermatology, Huashan Hospital, Shanghai Medical College, Fudan University, Shanghai, People’s Republic of China, 5 Shenzhen Key Laboratory for Translational Medicine of Dermatology, Shenzhen-PKU-HKUST Medical Center, Shenzhen, People’s Republic of China Abstract Background: JAK2 V617F, a somatic point mutation that leads to constitutive JAK2 phosphorylation and kinase activation, has been incorporated into the WHO classification and diagnostic criteria of myeloid neoplasms. Although various approaches such as restriction fragment length polymorphism, amplification refractory mutation system and real-time PCR have been developed for its detection, a generic rapid closed-tube method, which can be utilized on routine genetic testing instruments with stability and cost-efficiency, has not been described. Methodology/Principal Findings: Asymmetric PCR for detection of JAK2 V617F with a 39-blocked unlabeled probe, saturate dye and subsequent melting curve analysis was performed on a Rotor-GeneH