急性白血病MLL融合基因新检测方法的建立和应用
Establishment and Application of A Novel Method for Detecting MLL Fusion Genes of Acute Leukemia
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Date
2012-08Author
赵晓明
王航(厦门大学附属中山医院血液科)
李志鹏
鹿全意(厦门大学附属中山医院血液科)
李庆阁
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- 生命科学-已发表论文 [5877]
Abstract
[中文文摘]本研究旨在建立一种快速检测急性白血病中常见MLL融合基因的方法。针对10种最常见的MLL融合基因,首先通过检索文献及数据库,确定已报道的所有融合形式并据此设计引物和探针。其次,以构建的16个阳性质粒和阴性细胞系建立和优化多重实时PCR检测体系。最后,利用所收集的54份白血病标本对该体系进行临床评估,并对检出的阳性标本进行测序验证。结果:该体系可对所有阳性质粒进行有效检测,且灵敏度均可达10个拷贝。在54份标本中,检出MLL-AF4、MLL-AF9、MLL-AF10、MLL-ELL 4种类型的融合基因,对阳性标本进行测序,测序结果与检测结果一致。结论:本研究建立了一种筛查MLL融合基因的多重实时PCR方法,能够对发生频率约占MLL融合类型90%的10种融合基因进行检测。该体系具有快速、灵敏、特异、可靠的优点,将有助于临床上MLL融合基因阳性患者的诊断和管理。[英文文摘]This study was aimed to establish an efficient method to detect 10 common MLL fusion genes in patients with acute leukemia.Firstly,the relevant references and databases were searched to throughly inivestigate all fusion breakpoints;the primers and probes were designed according to nearly all the involved fusion types of gene.Then the multiplex real-time PCR system was established and optimized by using the established 16 positive plasmids and negative cell lines.Finally,the detection system was clinically evaluated by means of collected 54 samples of leukemia. The results indicated that the established detection system could efficiently detect all positive plasmids with sensibility to 10copies. Four kinds of fusion gene types such as MLL-AF4,MLL-AF9,MLL-AF10,MLL-ELL could be detected in 54 samples,the sequencing of positive samples showed consistency of sequencing results with detectionresults. It is concluded that a novel multiplex real-time PCR detection method is established which can detect 10 common MLL fusion genes covering about 90% of the cases harboring MLLfusions. This method is fast,sensitive,specific and reliable,and should be an useful clinical tool for identification and management of leukemia patients with MLLfusions.