A new high-throughput sequencing method for determining diversity and similarity of T cell receptor (TCR) α and β repertoires and identifying potential new invariant TCR α chains.
A new high-throughput sequencing method for determining diversity and similarity of T cell receptor (TCR) α and β repertoires and identifying potential new invariant TCR α chains.
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一种新的高通量测序方法，用于确定T细胞受体（TCR）α和β曲目的多样性和相似性，并确定潜在的新不变TCRα链。
DOI：
10.1186/s12865-016-0177-5
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发表时间：
2016-10-11
期刊：
影响因子：
3
通讯作者：
Suzuki R
中科院分区：
文献类型：
--
作者：
Kitaura K;Shini T;Matsutani T;Suzuki R
T细胞受体（TCR）基因的高通量测序是分析T淋巴细胞抗原特异性、克隆性和多样性的有力工具。在这里，我们开发了一种新的TCR库分析方法，使用454 DNA测序技术结合衔接子连接介导的聚合酶链反应（PCR）。该方法允许扩增所有TCR基因而没有PCR偏倚。为了比较个体间表达的TCR库的基因使用、多样性和相似性，我们对来自20名健康人个体的外周血单核细胞中的TRA和TRB基因进行了下一代测序（NGS）。从来自20个个体的总共267，037个序列读段中，鉴定了149，216个独特序列读段。观察到几个V和J基因的优先使用，而TRAV与TRAJ的一些重组似乎受到限制。TCR的多样性程度在TRA和TRB之间没有显著差异，而TRA谱系在个体之间比TRB谱系更相似。TRA的个体间相似性很大程度上取决于两个或多个个体之间频繁存在的共享TCR。可公开获得的TRA具有近种系TCR，其具有较短的CDR 3。值得注意的是，共享的TRA序列，特别是那些在大量个体中共享的TRA序列，通常包含与来自恒定自然杀伤T细胞和粘膜相关恒定T细胞的恒定TCRα相关的TCRα。这些结果表明，通过NGS检索共享TCR将有助于鉴定潜在的新的恒定TCRα链。这种NGS方法将能够在克隆水平上对TCR库进行全面的定量分析。本文的在线版本（doi：10.1186/s12865-016-0177-5）包含补充材料，可供授权用户使用。
High-throughput sequencing of T cell receptor (TCR) genes is a powerful tool for analyses of antigen specificity, clonality and diversity of T lymphocytes. Here, we developed a new TCR repertoire analysis method using 454 DNA sequencing technology in combination with an adaptor-ligation mediated polymerase chain reaction (PCR). This method allows the amplification of all TCR genes without PCR bias. To compare gene usage, diversity and similarity of expressed TCR repertoires among individuals, we conducted next-generation sequencing (NGS) of TRA and TRB genes in peripheral blood mononuclear cells from 20 healthy human individuals. From a total of 267,037 sequence reads from 20 individuals, 149,216 unique sequence reads were identified. Preferential usage of several V and J genes were observed while some recombinations of TRAV with TRAJ appeared to be restricted. The extent of TCR diversity was not significantly different between TRA and TRB, while TRA repertoires were more similar between individuals than TRB repertoires were. The interindividual similarity of TRA depended largely on the frequent presence of shared TCRs among two or more individuals. A publicly available TRA had a near-germline TCR with a shorter CDR3. Notably, shared TRA sequences, especially those shared among a large number of individuals’, often contained TCRα related with invariant TCRα derived from invariant natural killer T cells and mucosal-associated invariant T cells. These results suggest that retrieval of shared TCRs by NGS would be useful for the identification of potential new invariant TCRα chains. This NGS method will enable the comprehensive quantitative analysis of TCR repertoires at a clonal level. The online version of this article (doi:10.1186/s12865-016-0177-5) contains supplementary material, which is available to authorized users.
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影响因子：
9.8
作者：
Martin E;Treiner E;Duban L;Guerri L;Laude H;Toly C;Premel V;Devys A;Moura IC;Tilloy F;Cherif S;Vera G;Latour S;Soudais C;Lantz O
通讯作者：
Lantz O
影响因子：
56.9
作者：
Arstila, TP;Casrouge, A;Kourilsky, P
通讯作者：
Kourilsky, P
DOI：
10.1073/pnas.85.23.8998
发表时间：
1988-12-01
影响因子：
11.1
作者：
FROHMAN, MA;DUSH, MK;MARTIN, GR
通讯作者：
MARTIN, GR
影响因子：
4.4
作者：
Huang, CY;Kanagawa, O
通讯作者：
Kanagawa, O
影响因子：
7
作者：
Freeman, J. Douglas;Warren, Rene L.;Holt, Robert A.
通讯作者：
Holt, Robert A.