scRNAseq Dataset of lungs from WT and Clec1a KO mice following primary or secondary E. coli lung infection

Single-cell RNA sequencing (scRNAseq) sample preparationWT and Clec1a mice were infected intracheally with E. coli and lungs were recovered and dissociated at day 1, 3 or 7 post-infection. The obtained cell suspension was FACS-sorted for CD45+, CD45+Ly6G- and CD45-CD31+ cell populations (BD FACS ARIA, BD Biosciences). Then, in order to reduce the predominant neutrophil population and to enrich in endothelial cells for scRNAseq experiments, purified cells were mixed with as follow (6·105 CD45+ Ly6G- cells, 3·105 CD45+ cells and 105 CD45- CD31+ cells). ScRNAseq library preparation and sequencingCells were incubated with TotalSeq barcoded-antibodies for each sample, in order to identify the origin of the cell after the RNA library creation. Around 16000 cells per reaction were then processed with a 10X Chromium Next Controller following the 3’ V3 protocol. RNA libraries were then prepared with the Chromium Single Cell 3′ Library and Gel Bead Kit v2 (PN-120267) and Chromium Next GEM Single Cell 3′ Library and Gel Bead Kit v3.1 (PN-1000121) (10X Genomics). Libraries were then sequenced with an Illumina NovaSeq 6000. scRNAseq data analysisSequenced data underwent the Cellranger-7.2.0 pipeline, in order to perform sample demultiplexing, sequence alignment, barcode processing and unique molecular identity (UMI) counting. Filtered matrixes were processed and cells containing more than 5% of mitochondrial genes were cut off. Dataset integration, and clustering was performed using the Seurat package (v4.4.0). The uploaded data correspond to the seurat objects containing the integrated datasets (with cell annotation and clustering), for primary and secondary infection (alldata.Primary and alldata.secondary). Furthermore, filtered matrixes used to generate these Seurat objects (Primary Infection scRNAseq Filtered Matrix Part 1-3, Secondary Infection scRNAseq Filtered Matrix) were provided.单细胞RNA测序（single-cell RNA sequencing, scRNAseq）样本制备：对野生型（WT）与Clec1a小鼠经气管内感染大肠杆菌（E. coli），于感染后第1、3、7天采集肺组织并解离。将获取的细胞悬液通过流式细胞术（FACS，BD FACS ARIA，BD Biosciences）分选出CD45+、CD45+Ly6G-及CD45-CD31+细胞群。为减少优势中性粒细胞群体并富集内皮细胞以用于scRNAseq实验，按如下比例混合纯化后的细胞：6×10^5个CD45+Ly6G-细胞、3×10^5个CD45+细胞及1×10^5个CD45-CD31+细胞。 scRNAseq文库制备与测序：对每个样本，将细胞与TotalSeq条形码抗体进行孵育，以便在RNA文库构建完成后识别细胞来源。每个反应体系中约含16000个细胞，随后使用10X Chromium Next Controller并遵循3’ V3试剂盒流程进行处理。随后使用Chromium Single Cell 3′ Library and Gel Bead Kit v2（PN-120267）和Chromium Next GEM Single Cell 3′ Library and Gel Bead Kit v3.1（PN-1000121，10X Genomics）制备RNA文库。最终使用Illumina NovaSeq 6000测序平台完成文库测序。 scRNAseq数据分析：测序数据采用Cellranger-7.2.0分析流程，完成样本解复用、序列比对、条形码处理与唯一分子标识符（unique molecular identity, UMI）计数。对过滤后的表达矩阵进行处理，剔除线粒体基因占比超过5%的细胞。数据集整合与细胞聚类分析采用Seurat包（v4.4.0）完成。 本次上传的数据包含针对原发感染与继发感染的整合数据集Seurat对象（已完成细胞注释与聚类分析，分别对应alldata.Primary与alldata.secondary）。此外，还提供了用于生成上述Seurat对象的过滤表达矩阵，包括原发感染scRNAseq过滤表达矩阵第1-3部分，以及继发感染scRNAseq过滤表达矩阵。