1月龄Balb/c小鼠胸腺、脾脏、肝脏、小肠和血液TRBV-TRBJ1-1 CDR3谱型分析

目的初步分析1月龄Balb/c小鼠胸腺、脾脏、肝脏、小肠和血液中TRBV-TRBJ1-1 CDR3谱型特征。方法提取1月龄Balb/c小鼠胸腺、脾脏、肝脏、小肠和血液样本的总RNA,逆转录成cDNA,以22条TRBV基因家族为上游引物,共同的TRBJ1-1基因为下游引物(荧光标记),PCR扩增包含完整CDR3区片断,毛细管电泳激光DNA扫描技术分析CDR3谱型的长度和多态性。结果胸腺样本毛细管电泳激光扫描图呈现标准的中间高两端低的多态性分布;各家族CDR3区最长和最短比较,多数为相差8～15个氨基酸的谱型分布。脾脏、肝脏、小肠、血液样本扫描图显示多数家族呈多态性分布,但均出现数量不等的单峰、寡峰、偏峰、低峰;多数家族表现为相差3～13个氨基酸的谱型分布,部分家族在预测大小位置无对应峰型。结论 1月龄Balb/c小鼠胸腺中基本包含TCR基因随机重排的多种克隆性T细胞;脾脏、肝脏、小肠、血液表现为单、寡、多克隆形式的T细胞分布,部分家族T细胞缺失,本实验结果可为进一步研究Balb/c小鼠的T细胞发育、分布、应答及CDR3谱型与疾病关系提供方法和基础。

Analysis of the TRBV-TRBJ1-1 CDR3 repertoire in the thymus,spleen,liver,small intestine,and blood of one-month-old Balb/c mouse

This study aims to primarily analyze the TRBV-TRBJ1-1 CDR3 repertoire in the thymus,spleen,liver,small intestine and blood of one-month-old Balb/c mouse.Firstly,total RNA was extracted from the thymus,spleen,liver,small intestine and blood of one-month-old Balb/c mouse and reverse transcribed into cDNA.PCR was used to amplify the TRBV-TRBJ1-1 CDR3 region with the primers;capillary electrophoresis DNA scan was used to analysis of the CDR3 length repertoire and the diversity.In the thymus,the CDR3 spectratyping of TRBV families showed a standard diversity distribution;the pattern of some TRBV families showed normal distribution and there were 8-15 amino acid discrepancies between the longest CDR3 and shortest CDR3.In the liver,small intestine,and blood,most of families presented diversity distribution,but there were different number of mono-peak/oligo-peak/poly-peak/low-peak,the pattern showed non-normal distribution and the discrepancies were 3-13 amino acid.Portion of families showed no corresponding peak in the prediction position.The study suggested that,the thymus of one-month-old Balb/c mouse contains varied clonal T cells having random rearrange TCR genes;while in the spleen,liver,small intestine and blood,monoclonal/oligoclonal/polyclonal of T cells are presented and part of families were defected.The experimental results may be helpful to provide methods and basis for further study of the development,distribution,response of T cell and CDR3 repertoire in relation to diseases in Balb/c mouse.