摘要
本研究旨在建立检测急性髓系细胞白血病(AML)患者NPM1基因突变的方法。针对NPM1基因突变集中区域设计引物/探针,建立高分辨熔解曲线方法(HRM)和等位基因特异PCR方法(AS-PCR),通过89份AML标本进行了临床评估,并采用毛细管电泳法(CE)和测序法作为对照进行了验证。结果表明,通过上述4种方法共检出阳性标本17例(19.1%),其中A型突变15例,B型和Nm型突变各1例。上述方法中HRM法检测全面,灵敏度为5%,而AS-PCR法有一定的漏检,但灵敏度高达0.01%。结论:考虑到操作的难易程度,同时也结合临床样品的检测情况,HRM在临床上可法用于NPM1突变筛查,而AS-PCR法可用于后续的微小残留病定量检测。
The purpose of this study was to establish real-time based methods for detection of NPM1 gene mutation in acute myeloid leukemia （AML）. Primers/probes were designed according to the clustered region of NPM1 mutations on exon 12. Two real-time PCR assays, including high resolution melting curve （HRM） and allele-specific PCR （AS- PCR）, were developed and clinically evaluated with 89 AML samples, which were parallelly detected by capillary electrophoresis （CE） and sequencing. The results showed that a total of 17 mutation-positive samples were detected, including type A （ 15 cases）, type B （ 1 case） and type Nm （ 1 case）. HRM assay could detect all mutant types, and the analytical sensitivity was around 5 %. In contrast, AS-PCR assay detected only 95 % mutant types, but its sensitivity was as high as 0. 01%. It is concluded that considering the characteristics of each method as well as the clinical evaluation results, HRM may be used for screening of NPM1 mutations at diagnosis, while the AS-PCR can be used for the MRD quantification during follow-up.
出处
《中国实验血液学杂志》
CAS
CSCD
2011年第4期999-1004,共6页
Journal of Experimental Hematology
