High-throughput Ig and TCR repertoire single-cell sequencing analysis in rhesus macaques

Recent advancements in microfluidics and high-throughput sequencing technologies have enabled recovery of paired heavy- and light- chain of immunoglobulins (Ig) and VDJ- and VJ- chains of T cell receptors (TCR) from thousands of single cells simultaneously. Due to the complexity of these polyclonal receptors, for many species single-cell immune repertoire sequencing assays are not yet commercially available. Rhesus macaques are one of the most well-studied model organisms of the human adaptive immune response; application of these new immune repertoire sequencing assays is highly relevant to vaccine and infectious disease studies. Here we use custom designed primers to target and enrich for every known Ig and TCR chain and isotype in the rhesus macaque animal model. We sequenced more than 110,000 cell barcodes from rhesus macaque repertoires using PBMC, splenocyte, and FACS-sorted T and B cell. We were able to recover every Ig and TCR isotype, measure clonal expansion in proliferating T cells, and pair repertoires with gene expression profiles of single cells. Our results establish the ability to perform single-cell based immune repertoire analysis in rhesus macaque. High-throughput single-cell sequencing of T and B cell receptor sequences from 6 samples (2 PBMC, splenocyte, stimulated B cells, and stimulated T cells divided into proliferating and non-proliferating fractions). In addition, 5' GEX profiles were also sequenced for all but one PBMC sample. This dataset includes the 5' GEM profiles.

微流控（microfluidics）与高通量测序技术的最新进展，使得我们能够同时从数千个单细胞中获取免疫球蛋白（immunoglobulin, Ig）的重链与轻链，以及T细胞受体（T cell receptor, TCR）的VDJ链与VJ链。由于此类多克隆受体结构复杂，目前针对多数物种的单细胞免疫组库测序检测技术仍未实现商业化应用。恒河猴是目前研究最为深入的人类适应性免疫应答模式生物之一，此类新型免疫组库测序检测技术的应用与疫苗及传染病研究高度相关。本研究通过定制设计的引物，靶向富集恒河猴模型中所有已知的Ig、TCR链及其同种型。我们对来自恒河猴免疫组库的超过11万个细胞条形码进行了测序，样本涵盖外周血单个核细胞（peripheral blood mononuclear cell, PBMC）、脾细胞，以及经荧光激活细胞分选（fluorescence-activated cell sorting, FACS）纯化的T细胞与B细胞。本研究成功获取了所有Ig与TCR同种型，实现了增殖性T细胞的克隆扩增定量，并将免疫组库数据与单细胞基因表达谱进行了联合分析。本研究结果证实，可在恒河猴模型中开展基于单细胞的免疫组库分析。本数据集包含6份样本的T、B细胞受体序列高通量单细胞测序数据，样本包括2份外周血单个核细胞样本、1份脾细胞样本、1份活化B细胞样本，以及1份活化T细胞样本（进一步分为增殖与非增殖两个组分）。此外，除1份外周血单个核细胞样本外，其余所有样本均完成了5'端基因表达（gene expression, GEX）谱测序。本数据集同时包含5'端微珠乳滴（Gel Bead in Emulsion, GEM）测序谱。