Files pertaining to data analyses performed and presented in the preprint, 'Porcine intestinal innate lymphoid cells and lymphocyte spatial context revealed through single-cell RNA sequencing' by Wiarda et al. 2022 are provided in this dataset. Single cell suspensions enriched for lymphocytes were obtained from ileum of two seven-week-old pigs and subjected to single-cell RNA sequencing (scRNA-seq). Peripheral blood mononuclear cells (PBMCs) were collected and processed for scRNA-seq in parallel. scRNA-seq was performed to provide transcriptomic profiles of lymphocytes in porcine ileum, with 31,983 cells annotated into 26 cell types. Deeper interrogation of data revealed previously undescribed cells in porcine intestine, including SELLhi γδ T cells, group 1 and group 3 innate lymphoid cells (ILCs), and four subsets of B cells. Single-cell transcriptomes in ileum were compared to those in porcine blood, and subsets of activated lymphocytes were detected in ileum but not periphery. Comparison to scRNA-seq human and murine ileum data revealed a general consensus of ileal lymphocytes across species. Lymphocyte spatial context in porcine ileum was conferred through differential tissue dissection prior to scRNA-seq. Antibody-secreting cells, B cells, follicular CD4 αβ T cells, and cycling T/ILCs were enriched in ileum with Peyer’s patches, while non-cycling γδ T, CD8 αβ T, and group 1 ILCs were enriched in ileum without Peyer’s patches. Data files included herein are .h5seurat files of the various cell subsets included in analyses of the manuscript. Files may be used to reconstruct different analyses and perform further data query. Scripts for original data analyses are found at https://github.com/USDA-FSEPRU/scRNAseq_Porcine_Ileum_PBMC. Raw data are available at GEO accession GSE196388. Data are available for online query at https://singlecell.broadinstitute.org/single_cell/study/SCP1921/intestinal-single-cell-atlas-reveals-novel-lymphocytes-in-pigs-with-similarities-to-human-cells. Resources in this dataset:Resource Title: Ileum_AllCells. File Name: Ileum_AllCells.tarResource Description: .h5seurat object of all the cells derived from ileum samples. Untar into .h5seurat file before use.Resource Title: GutBlood_IntegratedILCs. File Name: GutBlood_IntegratedILCs.tarResource Description: .h5seurat object of ILCs derived from both ileum and PBMC samples. Untar into .h5seurat file before use.Resource Title: Ileum_Bonly. File Name: Ileum_Bonly.tarResource Description: .h5seurat object of B cells and antibody-secreting cells derived from ileum samples. Untar into .h5seurat file before use.Resource Title: Ileum_CD4Tonly. File Name: Ileum_CD4Tonly.tarResource Description: .h5seurat object of non-naive CD4 ab T cells derived from ileum samples. Untar into .h5seurat file before use.Resource Title: Ileum_gdCD8Tonly. File Name: Ileum_gdCD8Tonly.tarResource Description: .h5seurat object of gd and CD8 ab T cells derived from ileum samples. Untar into .h5seurat file before use.Resource Title: Ileum_ILConly. File Name: Ileum_ILConly.tarResource Description: .h5seurat object of innate lymphoid cells (ILCs) derived from ileum samples. Untar into .h5seurat file before use.Resource Title: Ileum_MyeloidOnly. File Name: Ileum_MyeloidOnly.tarResource Description: .h5seurat object of myeloid lineage leukocytes derived from ileum samples. Untar into .h5seurat file before use.Resource Title: Ileum_NonImmuneOnly. File Name: Ileum_NonImmuneOnly.tarResource Description: .h5seurat object of non-immune cells derived from ileum samples. Untar into .h5seurat file before use.Resource Title: Ileum_TILConly. File Name: Ileum_TILConly.tarResource Description: .h5seurat object of all T cells and innate lymphoid cells (ILCs) derived from ileum samples. Untar into .h5seurat file before use.Resource Title: PBMC_AllCells. File Name: PBMC_AllCells.tarResource Description: .h5seurat object of all cells derived from PBMC samples. Untar into .h5seurat file before use.