多重RT-PCR联合毛细管电泳法检测BCR-ABL融合基因

doi:10.3969/j.issn.1673-8640.2014.03.006

检验医学 ›› 2014, Vol. 29 ›› Issue (3): 226-232.DOI: 10.3969/j.issn.1673-8640.2014.03.006

• 分子诊断技术的临床应用专题 • 上一篇下一篇

多重RT-PCR联合毛细管电泳法检测BCR-ABL融合基因

吕晓楠¹，叶辉铭^1，2，顾龙¹，曾骥孟¹

1. 厦门大学药学院转化医学中心，福建厦门 361005；2. 厦门大学附属中山医院临检中心，福建厦门 361004

收稿日期:2013-07-15 出版日期:2014-03-30 发布日期:2014-04-19
通讯作者: 曾骥孟，联系电话:0592-2187226。
作者简介:吕晓楠，女，1987年生，硕士，主要研究方向为分子诊断。
基金资助:
福建省自然科学基金计划资助项目(2012J01414)；厦门市科技局计划项目资助课题(3502Z20113013)

Detection of BCR-ABL fusion gene by multiplex RT-PCR combining with capillary electrophoresis

LÜ Xiaonan¹,YE Huiming^1,2,GU Long¹,ZENG Jimeng

1. Translational Medicine Research Center，School of Pharmaceutical Science，Xiamen University，Fujian Xiamen 361005，China；2.Department of Clinical Laboratory，Zhongshan Hospital，Xiamen University，Fujian Xiamen 361004，China

Received:2013-07-15 Online:2014-03-30 Published:2014-04-19

摘要/Abstract

摘要：

目的　建立多重逆转录-聚合酶链反应(RT-PCR)联合毛细管电泳法检测BCR-ABL融合基因，以期应用于慢性粒细胞白血病(CML)辅助诊断。方法　采用重叠延伸法构建BCR-ABL融合基因阳性模板，设计并优化检测BCR-ABL融合基因的多重RT-PCR联合毛细管电泳体系，对检测体系进行初步的性能评估。结果　多重RT-PCR技术检测BCR-ABL融合基因(e1a2、e13a2和e14a2的最低检测限在10²～10³拷贝/μL；50例临床确诊CML的患者应用该法检测BCR-ABL融合基因的阳性率为86.0%，其中e13a2型（20.0%），e14a2型（66.0%）；本方法与骨髓细胞培养染色体核型分析相比，阳性符合率为97.6%，阴性符合率为75.0%，总符合率为94.0%，2种方法具有较高的一致性（Kappa=0.765，P>0.05）。结论　成功建立检测BCR-ABL融合基因的多重RT-PCR联合毛细管电泳方法。该方法操作简单快捷、特异性好、灵敏度高，适合于临床上CML的辅助诊断和分子分型。

关键词: BCR-ABL融合基因, 慢性粒细胞白血病, 多重RT-PCR, 毛细管电泳

Abstract:

Objective　To establish a method for detecting BCR-ABL fusion gene by multiplex reverse transcription polymerase chain reaction ( RT-PCR),and to provide a useful tool for chronic myeloid leukemia (CML) auxiliary diagnosis. Methods　The construction of BCR-ABL fusion gene as positive template was made by overlap extension method. The method of multiplex RT-PCR combining with capillary electrophoresis was designed and optimized. Its primary performance was evaluated. Results　The lower detection limit for BCR-ABL fusion gene (ela2,e13a2 and e14a2) by multiplex RT-PCR was 10²-10³ copies/μL. The positive rate of this method to detect 50 CML patients was 86.0%(e13a2: 20.0% and e14a2: 66.0%). Compared with the results of chromosome karyotype analysis,the positive coincidence rate was 97.6%,and the negative coincidence rate was 75.0%. The 2 methods have high consistency (Kappa=0.765,P>0.05). Conclusions　The method of multiplex RT-PCR combining with capillary electrophoresis to detect BCR-ABL fusion gene is simple and rapid with good specificity and sensitivity and is very suitable for the clinical detection of CML.

Key words: BCR-ABL fusion gene, Chronic myeloid leukemia, Multiplex reverse transcription polymerase chain reaction, Capillary electrophoresis

中图分类号:

Q503

吕晓楠，叶辉铭，顾龙，曾骥孟. 多重RT-PCR联合毛细管电泳法检测BCR-ABL融合基因[J]. 检验医学, 2014, 29(3): 226-232.

Lü Xiaonan，YE Huiming，GU Long，ZENG Jimeng. Detection of BCR-ABL fusion gene by multiplex RT-PCR combining with capillary electrophoresis[J]. , 2014, 29(3): 226-232.

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多重RT-PCR联合毛细管电泳法检测BCR-ABL融合基因

Detection of BCR-ABL fusion gene by multiplex RT-PCR combining with capillary electrophoresis

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