正常人外周血αβ T细胞TCR β链CDR3多态性和长度分析  

Analysis of Polymorphism and Length of TCR beta Chain CDR3 Spectratyping in Peripheral Blood αβ T Cells of Healthy Donors

作　　者：胡云良[1] 杨介钻[2] 倪莉 姚新生[3] 

机构地区：[1]温州医学院附属第二医院检验科,浙江温州325027 [2]浙江大学医学院附属第一医院传内科,浙江杭州310003 [3]遵义医学院免疫学研究室,贵州遵义563003

出　　处：《中国生物化学与分子生物学报》2007年第6期492-498,共7页Chinese Journal of Biochemistry and Molecular Biology

摘　　要：采用免疫扫描谱型分析技术,分析正常人外周血T细胞TCRβ链CDR3的多态性和长度分布.提取6例正常人外周血单个核细胞(peripheral blood mononuclear cell-PBMC)的总RNA,逆转录成cDNA,以24个TCR BV基因家族为上游引物,共同的TCR BC基因为下游引物(荧光标记),6例样品PCR产物的基因扫描分析表明,24个TCRBV家族的CDR3谱型分析在1.5%琼脂糖凝胶电泳图上显示1个模糊条带.序列测定结果显示,各TCR BV家族的CDR3谱型均超过8个条带.GeneScan分析证明,正常人外周血T细胞的24个TCR BV家族的CDR3谱型为标准的高斯分布,各家族的CDR3表达频率相近,呈现不同的CDR3多态性和不同的CDR3长度.多数TCR BV家族产物为框架内重排(相邻2个PCR产物相差3个碱基),少数TCR BV家族表现为两个峰群的重排.该研究对正常人TCRβ链CDR3谱系的详细分析将为T细胞应答、TCR重组等研究提供基础.利用免疫谱型分析技术能监测到TCR CDR3谱型分布特征和表达频率的变化.The objective of this study was to analyze the polymorphism and length of the TCR beta chain CDR3 spectratyping of peripheral blood αβ T cells from healthy donors by immunoscope spectratyping technique. The total RNA of peripheral blood mononuclear cell （PBMC） from 6 healthy donors was transcripted reversely into eDNA. The eDNA of TCR CDR3 was amplified by PCR technique. The products of PCR were analyzed with sequencing gel and gene-scanning techniques respectively. The PCR products exhibited an obscure band on 1.5 % agarose gel electrophoresis, and each TCR BV family exhibited more than 8 bands on 6% sequencing gel electrophoresis. The CDR3 spectratyping of 24 TCR BV families showed a standard Gaussian distribution and gave different CDR3 polymorphisms and lengths and they showed also similar expression frequency of CDR3 among the families. Most of CDR3 in TCR BV families recombined in the frame（there was 3 bp discrepancy between two products）while some of the CDR3 showed two groups of gene rearrangement in the 6 healthy donors. The study suggested that the detailed analysis of the repertoires of CDR3 gene of TCR beta chain will lay the foundation for the further study on T cell response and TCR recombination. The characters of spectratyping distribution and changes of expression frequency of TCR CDR3 can be monitored by immunoscope spectratyping technique.

关 键 词：T淋巴细胞受体 互补决定区3 免疫扫描谱型分析 基因扫描 

分 类 号：R392.12[医药卫生—免疫学]