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目的采用二代测序技术监测复发儿童B细胞急性淋巴白血病(B cell acute lymphoblastic leukemia,B-ALL)的克隆演变。方法 20例B-ALL患儿的初诊和复发骨髓标本,分别提取全基因组DNA,通过多重PCR方法扩增免疫球蛋白重链基因(immunoglobulin heavy chain,IGH)互补决定区3(complementarity determining region 3,CDR3)序列并进行二代测序检测。结果 20例B-ALL患儿初诊骨髓标本IGH CDR3测序结果显示,17例有CDR3特异性序列,其中9例有1种CDR3特异性序列,7例有2种CDR3特异性序列,1例有3种CDR3特异性序列;复发骨髓标本IGH CDR3测序结果显示,16例有CDR3特异性序列,其中9例有1种CDR3特异性序列,7例有2种CDR3特异性序列;对初诊和复发标本CDR3序列进行比对分析,发现4例患儿出现克隆演变。结论采用二代测序技术可监测儿童B-ALL复发过程中的克隆演变。
Objective To monitor the clonal evolution of childhood B cell acute lymphoblastic leukemia (B-ALL) using second-generation sequencing. Methods Twenty cases of newly diagnosed and recurrent bone marrow samples of B-ALL children were enrolled in this study. Genomic DNA was extracted from each group. Multiplex PCR was used to amplify the complementarity determining region 3 (IGH) 3 of the immunoglobulin heavy chain (IGH) CDR3) sequence and second-generation sequencing. Results The results of IGH CDR3 sequencing in 20 cases of newly diagnosed B-ALL patients showed that there were 17 CDR3-specific sequences in 9 cases, 1 CDR3-specific sequence in 9 cases, 2 CDR3-specific sequences in 7 cases and 3 cases in 3 cases The sequence of IGH CDR3 in the recurrent bone marrow specimens showed that there were 16 CDR3-specific sequences in 9 cases, 1 CDR3-specific sequence in 9 cases and 2 CDR3-specific sequences in 7 cases. Sequence analysis of the alignment showed that 4 cases of children with clonal evolution. Conclusion The second generation sequencing technique can be used to monitor the clonal evolution of childhood B-ALL relapse.