Retrieve the fastq files from SRA and then align to the canine genome. Instructions to download the fastq files can be found in 📁 input. The alignment code is currently not provided, but can be shared if desired. If you want it, you can request through GitHub.
Download the supplementary zip folder on the NCBI GEO project page as decribed in 📁 input directory. In addition to the count matrices you will also need the .csv files located in the 📁 sheets directory. Once all files are in place, you will need the runScript_pbmc.R which contains the analysis code and you will have to source the customFunctions.R.
NOTE: the file paths in runScript_pbmc.R will likely need to be modified when reading/writing data to run on your system.
Download the supplementary .rds file on the NCBI GEO project page as described in 📁 input directory. There is an .rds file for just the healthy dogs, combined dataset, and a file for each major immune cell subset. These files can be loaded into runScript_pbmc.R to bypass some of the more computationally intense steps, or they can be used for your own exploration!
This work was completed on the UC boulder Alpine supercomputer on a Linux operating system. All code was run in a conda environment and I am happy to share the .yml file upon request.
Otherwise, see below for packages loaded into the system.
> sessionInfo()
R version 4.1.1 (2021-08-10)
Platform: x86_64-conda-linux-gnu (64-bit)
Running under: Red Hat Enterprise Linux 8.4 (Ootpa)
Matrix products: default
BLAS/LAPACK: /projects/dyammons@colostate.edu/software/anaconda/envs/r_env/lib/libopenblasp-r0.3.18.so
locale:
[1] LC_CTYPE=C.UTF-8 LC_NUMERIC=C LC_TIME=C
[4] LC_COLLATE=C LC_MONETARY=C LC_MESSAGES=C
[7] LC_PAPER=C LC_NAME=C LC_ADDRESS=C
[10] LC_TELEPHONE=C LC_MEASUREMENT=C LC_IDENTIFICATION=C
attached base packages:
[1] stats4 stats graphics grDevices utils datasets methods
[8] base
other attached packages:
[1] ggtree_3.2.1 ape_5.6-2
[3] scuttle_1.4.0 scRNAseq_2.8.0
[5] ggpubr_0.4.0 slingshot_2.2.1
[7] TrajectoryUtils_1.2.0 SingleCellExperiment_1.16.0
[9] princurve_2.1.6 clusterProfiler_4.2.2
[11] msigdbr_7.5.1 ggsankey_0.0.99999
[13] lemon_0.4.5 reshape_0.8.9
[15] viridis_0.6.2 viridisLite_0.4.1
[17] SingleR_1.8.1 SeuratDisk_0.0.0.9019
[19] RColorBrewer_1.1-3 pheatmap_1.0.12
[21] DESeq2_1.34.0 SummarizedExperiment_1.24.0
[23] Biobase_2.54.0 MatrixGenerics_1.6.0
[25] matrixStats_0.62.0 GenomicRanges_1.46.1
[27] GenomeInfoDb_1.30.1 IRanges_2.28.0
[29] S4Vectors_0.32.2 BiocGenerics_0.40.0
[31] colorspace_2.0-3 ggrepel_0.9.1
[33] cowplot_1.1.1 scales_1.2.1
[35] patchwork_1.1.2 DoubletFinder_2.0.3
[37] clustree_0.4.4 ggraph_2.0.5
[39] forcats_0.5.2 stringr_1.4.1
[41] dplyr_1.0.10 purrr_0.3.5
[43] readr_2.1.2 tidyr_1.2.1
[45] tibble_3.1.8 ggplot2_3.3.6
[47] tidyverse_1.3.1 SeuratObject_4.1.3
[49] Seurat_4.3.0
loaded via a namespace (and not attached):
[1] rappdirs_0.3.3 rtracklayer_1.54.0
[3] scattermore_0.8 bit64_4.0.5
[5] knitr_1.40 irlba_2.3.5
[7] DelayedArray_0.20.0 data.table_1.14.4
[9] AnnotationFilter_1.18.0 KEGGREST_1.34.0
[11] RCurl_1.98-1.6 generics_0.1.3
[13] GenomicFeatures_1.46.5 ScaledMatrix_1.2.0
[15] RSQLite_2.2.18 shadowtext_0.1.1
[17] RANN_2.6.1 future_1.29.0
[19] bit_4.0.4 tzdb_0.3.0
[21] enrichplot_1.14.2 spatstat.data_3.0-0
[23] xml2_1.3.3 lubridate_1.9.0
[25] httpuv_1.6.5 assertthat_0.2.1
[27] xfun_0.34 hms_1.1.2
[29] babelgene_22.9 promises_1.2.0.1
[31] restfulr_0.0.15 progress_1.2.2
[33] fansi_1.0.3 dbplyr_2.1.1
[35] readxl_1.4.1 igraph_1.2.11
[37] DBI_1.1.3 geneplotter_1.72.0
[39] htmlwidgets_1.5.4 spatstat.geom_3.0-5
[41] ellipsis_0.3.2 backports_1.4.1
[43] annotate_1.72.0 biomaRt_2.50.3
[45] deldir_1.0-6 sparseMatrixStats_1.6.0
[47] vctrs_0.5.1 ensembldb_2.18.3
[49] ROCR_1.0-11 abind_1.4-5
[51] cachem_1.0.6 withr_2.5.0
[53] ggforce_0.4.1 progressr_0.11.0
[55] sctransform_0.3.5 GenomicAlignments_1.30.0
[57] treeio_1.18.1 prettyunits_1.1.1
[59] goftest_1.2-3 cluster_2.1.4
[61] DOSE_3.20.1 ExperimentHub_2.2.1
[63] lazyeval_0.2.2 crayon_1.5.2
[65] genefilter_1.76.0 hdf5r_1.3.7
[67] spatstat.explore_3.0-5 labeling_0.4.2
[69] pkgconfig_2.0.3 tweenr_2.0.2
[71] ProtGenerics_1.26.0 nlme_3.1-160
[73] rlang_1.0.6 globals_0.16.1
[75] lifecycle_1.0.3 miniUI_0.1.1.1
[77] downloader_0.4 filelock_1.0.2
[79] BiocFileCache_2.2.1 modelr_0.1.8
[81] rsvd_1.0.5 AnnotationHub_3.2.2
[83] cellranger_1.1.0 polyclip_1.10-4
[85] lmtest_0.9-40 Matrix_1.5-1
[87] aplot_0.1.2 carData_3.0-5
[89] zoo_1.8-9 reprex_2.0.1
[91] ggridges_0.5.4 rjson_0.2.21
[93] png_0.1-7 bitops_1.0-7
[95] KernSmooth_2.23-20 Biostrings_2.62.0
[97] blob_1.2.3 DelayedMatrixStats_1.16.0
[99] qvalue_2.26.0 parallelly_1.32.1
[101] spatstat.random_3.1-3 rstatix_0.7.0
[103] gridGraphics_0.5-1 ggsignif_0.6.4
[105] beachmat_2.10.0 memoise_2.0.1
[107] magrittr_2.0.2 plyr_1.8.7
[109] ica_1.0-3 zlibbioc_1.40.0
[111] scatterpie_0.1.7 compiler_4.1.1
[113] BiocIO_1.4.0 fitdistrplus_1.1-8
[115] Rsamtools_2.10.0 cli_3.6.0
[117] XVector_0.34.0 listenv_0.8.0
[119] pbapply_1.5-0 MASS_7.3-58.1
[121] tidyselect_1.2.0 stringi_1.7.8
[123] yaml_2.3.6 GOSemSim_2.20.0
[125] BiocSingular_1.10.0 locfit_1.5-9.6
[127] grid_4.1.1 fastmatch_1.1-3
[129] tools_4.1.1 timechange_0.1.1
[131] future.apply_1.10.0 parallel_4.1.1
[133] rstudioapi_0.14 gridExtra_2.3
[135] farver_2.1.1 Rtsne_0.16
[137] BiocManager_1.30.19 digest_0.6.30
[139] shiny_1.7.1 Rcpp_1.0.9
[141] car_3.1-0 broom_1.0.1
[143] BiocVersion_3.14.0 later_1.3.0
[145] RcppAnnoy_0.0.20 httr_1.4.4
[147] AnnotationDbi_1.56.2 rvest_1.0.3
[149] XML_3.99-0.12 fs_1.5.2
[151] tensor_1.5 reticulate_1.26
[153] splines_4.1.1 uwot_0.1.14
[155] yulab.utils_0.0.5 tidytree_0.3.9
[157] spatstat.utils_3.0-1 graphlayouts_0.8.3
[159] sp_1.5-1 ggplotify_0.1.0
[161] plotly_4.10.1 xtable_1.8-4
[163] jsonlite_1.8.3 tidygraph_1.2.2
[165] ggfun_0.0.8 R6_2.5.1
[167] pillar_1.8.1 htmltools_0.5.3
[169] mime_0.12 glue_1.6.2
[171] fastmap_1.1.0 BiocParallel_1.28.3
[173] BiocNeighbors_1.12.0 interactiveDisplayBase_1.32.0
[175] codetools_0.2-18 fgsea_1.20.0
[177] utf8_1.2.2 lattice_0.20-45
[179] spatstat.sparse_3.0-0 curl_4.3.3
[181] leiden_0.4.3 GO.db_3.14.0
[183] limma_3.50.3 survival_3.4-0
[185] munsell_0.5.0 DO.db_2.9
[187] GenomeInfoDbData_1.2.7 haven_2.4.3
[189] reshape2_1.4.4 gtable_0.3.1