single cell RNA sequencing of spleen mesenchymal reticular cells

We report the single cell transcriptome of mouse splenic mesenchymal reticular cells obtained from Ccl19-cre x R26R-EYFP mice on a C57BL/6 background. Spleen CD45- CD31- Ter119- cells that expressed at least one of the markers CD157, PDPN and EYFP were flow cytometrically sorted and encapsulated into emulsion droplets using a Chromium Controller (10x Genomics) and libraries prepared. scRNA-seq libraries were constructed using Chromium Single Cell 3' v3 Reagent Kit according to the manufacturer's protocol. Cells were examined under a microscope and counted with a hemocytometer before loading. Libraries were run on the HiSeq2500 for 100 bp paired end Illumina sequencing.本研究报道了C57BL/6背景下Ccl19-cre x R26R-EYFP小鼠来源的脾脏间质网状细胞的单细胞转录组。实验中通过流式细胞术分选出脾脏中CD45⁻、CD31⁻、Ter119⁻且至少表达CD157、PDPN及EYFP其中一种标志物的细胞；使用Chromium控制器（10x Genomics）将分选得到的细胞包裹进微液滴并制备文库。依照厂商说明书，采用Chromium单细胞3'端v3试剂试剂盒构建单细胞RNA测序（scRNA-seq）文库。上样前，通过显微镜观测细胞状态并使用血细胞计数板完成细胞计数。最终将文库置于HiSeq2500测序仪上进行100 bp双端Illumina测序。