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目的:采用不同方法检测慢性骨髓增殖性疾病(CMPD)患者的JAK2V617F突变率,评估临床应用的可能性。方法:提取84例Ph阴性CMPD患者单个核细胞DNA,利用RealtimePCR联合Taqman-MGB探针检测骨髓增殖性疾病患者的JAK2V6A7F的发生率,并利用限制性片段长度多态性(RFLP)对照检测CMPD患者的JAK2V617F突变率,用基因测序验证两种结果。结果:RealtimePCR联合Taqman-MGB探针检测84例Ph(-)CMPD患者,42例存在JAK2V617F,突变率为50%;其中PV、ET、IMF患者的突变率分别为76.7%(23/30)、25.9%(7/27)、42.5%(9/20),经RFLP检测84例Ph(-)CMPD总的突变率为33.3%,测序结果验证了RealtimePCR联合Taqman-MGB探针检测结果。结论:Taqman-MGB探针联合RealtimePCR方法有较高的准确率,可以作为检测JAK2V617F突变的方法。
Objective: To detect the mutation rate of JAK2V617F in patients with chronic myeloproliferative disease (CMPD) by different methods and to evaluate the possibility of clinical application. Methods: The DNA of 84 patients with Ph-negative CMPD was extracted. Real-time PCR and Taqman-MGB probe were used to detect the incidence of JAK2V6A7F in patients with myeloproliferative diseases. The patients with CMPD were detected by restriction fragment length polymorphism (RFLP) The JAK2V617F mutation rate was verified by sequencing both results. Results: 84 patients with Ph (-) CMPD were detected by RealtimePCR and Taqman-MGB probe. 42 patients had JAK2V617F with a mutation rate of 50%. The mutation rates in patients with PV, ET and IMF were 76.7% (23/30) 25.9% (7/27) and 42.5% (9/20) respectively. The total mutation rate of Ph (-) CMPD in 84 cases detected by RFLP was 33.3%. The sequencing results validated RealtimePCR and Taqman-MGB probe. Conclusion: Taqman-MGB probe combined with RealtimePCR method has high accuracy and can be used as a method to detect JAK2V617F mutation.