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

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。

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.