LNA PCR检测骨髓增殖性疾病JAK2基因V617F突变

目的:建立一种检测酪氨酸激酶2(JAK2)基因V617F突变的锁核酸PCR(LNA PCR)方法,并探讨其对骨髓增殖性疾病(MPD)的临床应用价值。方法:收集68例确诊为MPD患者外周血或骨髓标本,其中真性红细胞增多症(PV)37例、原发性血小板增多症(ET)22例、原发性骨髓纤维化(MF)9例。根据JAK2基因V617F突变位点设计引物和LNA,建立快速检测JAK2基因V617F突变的LNA PCR方法,同时以DNA直接测序法进行验证,并检测上述标本JAK2基因V617F突变率。结果:所建立的LNA PCR方法能有效检测JAK2基因V617F突变,其特异性和灵敏度明显高于DNA测序法(P<0.05);LNA PCR方法可以检测出1%的JAK2基因V617F突变型杂合子。68例MPD患者标本共检测出JAK2基因V617F突变55例,检出率80.88%。其中37例PV中JAK2基因V617F突变34例(91.89%),22例ET中JAK2基因V617F突变15例(68.18%),9例MF中JAK2基因V617F突变6例(66.67%)。结论:LNA PCR方法检测灵敏度高,能有效提高JAK2基因V617F突变检出率。

LNA PCR Detection for V617F Mutation of JAK2 in Myeloproliferative Diseases

Objective: To develop a locked nucleic acid PCR( LNA PCR ) assay for detecting V617F mutation of JAK2 and investigate its clinical application value. Method: Specimens of 68 patients with myeloproliferative diseases(MPD) were collected from the department of hematology inpatient including 37 polycythemia vera (PV) cases, 22 essential thromboeythemia (ET) cases and 9 primary myelofibrosis (MF) cases. Design LNA for V617F mutation of JAK2 and develop LNA PCR assay were used to quickly detect V617F mutation of JAK2,than the resulte were proofed by DNA sequencing method. Results: The LNA PCR assay could effectively screen V617F mutation of JAK2. The detection sensitivity is 1% for JAK2 V617F mutation, and its specificity and sensitivity was significantly higher than that of the DNA sequencing method (P <0.05). All of 68 cases of MPD patients, 55 cases were found with the JAK2 gene V617F mutation, and the detection rate was 80.88%. Wherein, in 37 cases of patients with PV, the mutation was 91.89% (34/37), and in 22 cases of patients with ET, the mutation ratio rate was 68.18% (15/22), and in 9 cases of MF patients, the mutation ratio was 66.67% (6/9). Conclusion: The LNA PCR assay could detect the JAK2 V617F mutation with high sensitivity and thus lead to an increased detection rate.