CAJ | 학술논문

目的用改良的杂交探针结合溶解曲线新方法检测急性髓性白血病c-kit基因17号外显子上基因突变。方法利用Primer Premier v5.00、Tm Utility v1.3等软件设计引物探针，两个阶段探针分别针对D816、N820和N822等突变热点。取患者DNA样品5μl进行PCR扩增及熔解曲线分析，PCR扩增产物进行测序分析并与熔解曲线分析方法进行比较。结果阳性质粒灵敏度试验的结果显示， N820 G的灵敏度为10％，其余突变灵敏度均能达到5％。12例CBF-AML标本中，5例检测为阳性，突变率为41.7％，测序结果与测序检测结果相符。结论新方法具有方便快速、灵敏度高、特异性好等特点，可用于CBF-AML患者的个体化诊断和治疗。
목적용개량적잡교탐침결합용해곡선신방법검측급성수성백혈병c-kit기인17호외현자상기인돌변。방법이용Primer Premier v5.00、Tm Utility v1.3등연건설계인물탐침，량개계단탐침분별침대D816、N820화N822등돌변열점。취환자DNA양품5μl진행PCR확증급용해곡선분석，PCR확증산물진행측서분석병여용해곡선분석방법진행비교。결과양성질립령민도시험적결과현시， N820 G적령민도위10％，기여돌변령민도균능체도5％。12례CBF-AML표본중，5례검측위양성，돌변솔위41.7％，측서결과여측서검측결과상부。결론신방법구유방편쾌속、령민도고、특이성호등특점，가용우CBF-AML환자적개체화진단화치료。
Objective To detect c-kit gene mutations of acute myeloid leukemia in the hotspot exon 17 of the c-kit gene using a modified hybridization probe combining with melting curve .Methods Primer Premier v5.00 and Tm Utility v1.3 software were used to design the primers and probe , the probe contained two parts , the first segment of the probe could detect mutations around D 816 and the second segment could detect mutations at N 820 and N822.5 μl DNA samples from patients were used for PCR amplification and melting curve analysis .PCR products were sent to be sequenced,these results were compared with those of the mutation-positive plasmids.Results The detection sensitivity on positive control plasmids was 10%for N820 G and 5%for the other mutations .c-kit mutations were identified in 5 of 12 patients with CBF-AML ( 41.7%) , the result was confirmed by sequencing analysis . Conclusion The new method is simple,rapid,and specific with high sensitivity and could be used in individual diagnosis and treatment of CBF-AML patients.