利用Fine-tiling aCGH分析TCR基因重排鉴定T细胞白血病克隆  

Analysis of TCR gene rearrangement for identification of T cell leukemia clone by using Fine-tiling aCGH

作　　者：郑海涛[1] 叶铁真 陈少华[1] 杨力建[1] 卢育洪[1] 李扬秋[1,3] 

机构地区：[1]暨南大学医学院血液病研究所 [2]广州市妇女儿童医疗中心血液肿瘤科 [3]暨南大学再生医学教育部重点实验室,广东广州510632

出　　处：《细胞与分子免疫学杂志》2011年第5期483-486,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金资助项目(30871091);中央高校基本科研业务费专项资金资助项目(21610603)

摘　　要：目的:建立基于分析T细胞受体(TCR)基因重排而确定T细胞-急性淋巴细胞白血病(T-ALL)克隆的新方法,为研究T-ALL中涉及TCR基因位点的染色体易位提供基础。方法:提取1例T-ALL患者外周血单个核细胞(PBMC)的总DNA,利用精细定位的寡核苷酸阵列比较基因组杂交(fine-tiling aCGH)分析样本与对照组基因组DNA的差异,了解不同染色体上可能的断裂点和具体的位点,根据所提供的初步结果,从断裂点涉及的基因序列设计特异引物,采用连接介导PCR(LM-PCR)和序列分析等方法去找出与之发生重排的基因序列。并进一步通过RT-PCR检测TCR基因表达。结果:该例T-ALL患者fine-tiling aCGH结果显示14染色体TCRα/δ基因座出现4个断裂点,分别对应TCR Vδ1、Vδ2、Jδ1和Jδ2基因位点。通过LM-PCR、测序及序列分析,该例T-ALL患者的PBMC中TCR基因涉及Vδ1Dδ2Dδ3 Jδ1、Vδ2Dδ3 Jδ2重排。RT-PCR的结果也验证T-ALL表达该TCR基因重排。结论:结果表明,利用fine-tiling aCGH及LM-PCR技术可以找出TCR基因重排,并且该方法是可靠的,也是发现一些涉及TCR位点的融合基因的一条途径。AIM： To establish a new method which analyzes T cell receptor（TCR） gene rearrangement for identification of T cells acute lymphoblastic leukemia（T-ALL） clone,it will provide the basis for the study of T-ALL including the chromosome translocation involving TCR loci.METHODS： Total DNA was isolated from peripheral blood mononuclear cells（PBMC） of one case with T-ALL.Using the fine-tiling array comparative genomic hybridization（fine-tiling aCGH） to analyze the genomic DNA differences of the case and control group,we could find the breakpoints and their position in the chromosomes.According to the preliminary results,we could design the specific primers for the positions of the breakpoints relative to sequence.Furthermore,the ligation-mediated PCR（LM-PCR） and sequence analysis were used to identify the TCR gene rearrangement.And TCR gene expression was detected by RT-PCR.RESULTS： The fine-tiling aCGH results of the T-ALL showed that the TCRα/δ locus of chromosome 14 appeared four breakpoints,corresponding to TCR Vδ1,Vδ2,Jδ1 and Jδ2.By LM-PCR,sequencing and sequence analysis,TCR gene of the case of T-ALL was involved in Vδ1Dδ2Dδ3Jδ1,Vδ2Dδ3Jδ2 rearrangement.RT-PCR results also confirmed the expression of these TCR gene rearrangements.CONCLUSION： The results demonstrated that fine-tiling aCGH and LM-PCR techniques could be used to identify the TCR gene rearrangement as one of the best perfect methods.And it was also a way to find some fusion genes involving in TCR gene.

关 键 词：Fine-tilingaCGH LM-PCR TCR基因重排 

分 类 号：R392.11[医药卫生—免疫学]