The dynamics of immunoglobulin V-gene usage and clonotype expansion in mice after prime and boost immunizations as analyzed by NGS.
The dynamics of immunoglobulin V-gene usage and clonotype expansion in mice after prime and boost immunizations as analyzed by NGS.
复制标题
通过 NGS 分析小鼠初免和加强免疫后免疫球蛋白 V 基因使用和克隆型扩展的动态。
DOI：
10.1080/21645515.2017.1379638
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发表时间：
2017
影响因子：
4.8
通讯作者：
Lu,Shan
中科院分区：
文献类型：
--
作者：
Farfán-Arribas,DiegoJ;Liu,Shuying;Wang,Shixia;Lu,Shan
在本研究中，开发了一种改进的NGS方法来研究在仅包括两次DNA免疫的简单小鼠免疫模型中的B细胞库进化。5′RACE和Ion Torrent长读段的组合使得即使从少量外周小鼠血液也能够进行无偏倚的免疫球蛋白库分析。通过疫苗扩增的B细胞群体显示出相对强的克隆性。在用第一次疫苗剂量致敏后，我们观察到V区段基因和CDR 3使用的一致模式（公共特异性）。有趣的是，这种模式随着第二剂免疫而多样化-尽管接受了相同的疫苗方案，但在个体小鼠中相对不同（私人特异性）。然而，有几种情况下，在第一次接种后扩增的相同的公共V区段基因和CDR 3在第二次免疫后进一步扩增。总之，似乎通过疫苗接种扩增的主要克隆型最初是同质的亚群，其在第二剂量后多样化，导致不同小鼠中的不同“私有”克隆组成。这些结果建立了一个新的平台，有价值的进行纵向分析的IG种系基因的使用和克隆型进化在整个免疫方案中，在常用的动物模型。
In the current study, an improved NGS approach was developed to study the B-cell repertoire evolution in a simple mouse immunization model including only two DNA immunizations. The combination of 5′RACE and Ion Torrent long reads enabled unbiased immunoglobulin repertoire analysis even from small amounts of peripheral mouse blood. The B-cell population expanded by the vaccine displayed a relatively strong clonality. Upon priming with the first vaccine dose, we observed a consistent pattern of V-segment gene and CDR3 usage (public specificities). Interestingly, this pattern diversified with the second dose of immunization —it was relatively different in individual mice in spite of having received the same vaccine regimen (private specificities). Nevertheless, there were several instances in which the same public V-segment genes and CDR3s that were expanded after the first dose were further amplified after the second immunization. Taken together, it appears that the major clonotypes expanded by vaccination were originally a homogeneous subset that later diversified after a second dose leading to diverse “private” clonal compositions in different mice. These results established a new platform valuable to perform longitudinal analyses of the Ig germline gene usage and clonotype evolution throughout an immunization regimen in a commonly used animal model.