一步法多重RT-PCR对儿童急性淋巴细胞白血病常见融合基因的检测及意义

One-step multiplex RT-PCR for identifying common fusion transcripts in childhood acute lymphoblastic leukemia

摘要:

目的 探讨一步法多重RT-PCR 对儿童急性淋巴细胞白血病(ALL)常见4 种融合基因的检测效果。方法 2003 年1 月至2010 年12 月确诊的ALL 患儿76 例,采集所有患儿骨髓标本,提取细胞总RNA,一步法多重RT-PCR 或常规巢式PCR 法对E2A/PBX1、MLL/AF4、BCR/ABL 和TEL/AML1 融合基因进行检测;DNA测序对PCR 结果进行验证。结果 一步法多重RT-PCR 及DNA 测序验证显示76 例ALL 患儿中,TEL/AML1融合基因阳性12 例(产物长度分别为298 bp 9 例,259 bp 3 例), E2A/PBX1 融合基因阳性3 例(产物长度373 bp), BCR/ABL 融合基因阳性1 例(产物长度2124 bp),MLL/AF4 融合基因阳性7 例(产物长度分别为427 bp 1 例,673 bp 6 例),与常规巢式PCR 检出结果一致。结论 一步法多重RT-PCR 可应用于儿童ALL 常见融合基因的检测。

Abstract:

Objective To evaluate the efficiency of one-step multiplex RT-PCR for identifying four common fusion transcripts (TEL/AML1, E2A/PBX1, MLL/AF4 and BCR/ABL) in children with acute lymphoblastic leukemia (ALL). Methods Total RNA was extracted from bone marrow samples of 76 children who were newly diagnosed with ALL between January 2003 and December 2010. These RNAs were analyzed for TEL/AML1, E2A/PBX1, MLL/AF4 and BCR/ABL by one-step multiplex RT-PCR or common nested-multiplex PCR. The PCR products were confirmed by DNA sequencing. Results TEL/AML1 was found in 12 cases (the length of products was 298 bp in 9 cases and 259 bp in 3 cases), E2A/PBX1 was found in 3 cases (the length of products was 373 bp), BCR/ABL was found in 1 case (the length of products was 2 124 bp), and MLL/AF4 was found in 7 cases (the length of products was 427 bp in 1 case and 673 bp in 6 cases) using one-step multiplex RT-PCR combined with DNA sequencing. The results were consistent with those using common nested-multiplex PCR. Conclusions One-step multiplex RT-PCR may be another alternative for detection of common fusion transcripts in children with ALL.