高通量测序技术构建新型冠状病毒感染儿童T细胞受体β链免疫组库

Construction of TCRβ repertoires in children with SARS-CoV-2 infection using high-throughput sequencing technology

目的构建新型冠状病毒感染儿童T细胞受体(T cell receptor,TCR)β链免疫组库,分析新冠病毒感染患儿和健康儿童TCR免疫组库差异。方法收集新冠病毒Delta变异株感染儿童和健康对照儿童全血样本各五份,RNA质检和文库构建后,进行高通量测序,分析新冠病毒感染患儿和健康儿童TCR免疫组库克隆扩增和多样性指数的差异,统计TCRβ的VJ基因的使用频率,采用非配对T检验统计方法分析。结果成功利用高通量测序技术构建新冠病毒感染儿童TCRβ免疫组库,与健康儿童相比,新冠病毒感染儿童的TCR免疫组库多样性指数(9.78±1.23)显著低于健康儿童(13.40±2.12)(P<0.05),TCR克隆扩增指数(0.18±0.07)显著高于健康儿童(0.06±0.06)(P<0.05)。针对TCRβ免疫组库VJ基因使用频率初步比较发现,在新冠病毒感染儿童中,最常见的V基因和J基因分别是TRBV28和TRBJ2-1。结论利用高通量测序技术构建新冠病毒感染儿童TCRβ免疫组库,并发现新冠病毒感染儿童特征性的TCRβ免疫组库,阐述新冠病毒感染的T细胞免疫组库特征,对其他病毒感染后快速TCRβ免疫组库的构建具有重要参考意义。

Objective To construct T cell receptorβ(TCRβ)repertoires in children with SARS-CoV-2 infection,and analyze the differences in TCRβrepertoires between children with SARS-CoV-2 infection and healthy children.Methods Whole blood samples from 5 children infected with SARS-CoV-2 Delta variant and from 5 healthy children were collected.After RNA quality inspection and repertoires construction,high-throughput sequencing was conducted to analyze the differences in clonal expansion and diversity indices of the TCR repertoires between children infected with SARS-CoV-2 and healthy children.The frequency of use of TCRβVJ genes was statistically analyzed using unpaired T-tests.Results We successfully constructed the TCRβrepertoires of children infected with SARS-CoV-2 using high-throughput sequencing technology.The diversity index of the TCR repertoire in children infected with SARS-CoV-2(9.78±1.23)was significantly lower compared to that in healthy children(13.40±2.12)(P<0.05),and the TCR clonal expansion index in children infected with SARS-CoV-2(0.18±0.07)was significantly higher compared to that in healthy children(0.06±0.06)(P<0.05).A preliminary comparison of the frequency of use of TCRβrepertoire VJ genes found that,in children infected with the SARS-CoV-2,the most common V and J genes were TRBV28 and TRBJ2-1,respectively.Conclusions The construction of the TCRβrepertoires in children infected with SARS-CoV-2 using high-throughput sequencing technology has revealed characteristic features of the TCRβrepertoires in these children.This is of significant reference value for unveiling the characteristics of the T-cell repertoires in children infected with SARS-CoV-2 and for the rapid construction of TCRβimmunological repertoires in other viral infections.