split.by error in Clustered_DotPlot function

[apal6]

Hi,

Thank you for the package. I was able to use the function Clustered_DotPlot without using split.by parameter. I want to look at the expression at different timepoints and time2 column is there in my metadata but i get an error when I try to use split.by.

`Clustered_DotPlot(seurat_object = _Seurat, split.by = "time2", features = c("gene1"),

•               colors_use_exp = PurpleAndYellow())
  

Error in [.data.frame(percent_mat, , idx) : undefined columns selected`

Could you help?

Thank you!

[samuel-marsh]

Hi,

Are you only providing one gene to the function to plot? Could you paste the output of traceback() after the error occurs?

Thanks,
Sam

[samuel-marsh]

Hi @apal6,

Just wanted to followup on my previous message to try and figure out the issue.

Best,
Sam

[apal6]

Hi @samuel-marsh,

Thank you for following up.
No, I using multiple genes. Here is the output of traceback()

5: stop("undefined columns selected") 4: [.data.frame(percent_mat, , idx) 3: percent_mat[, idx] 2: Clustered_DotPlot_Multi_Group(seurat_object = seurat_object, features = features, split.by = split.by, colors_use_exp = colors_use_exp, exp_color_min = exp_color_min, exp_color_middle = exp_color_middle, exp_color_max = exp_color_max, exp_value_type = exp_value_type, print_exp_quantiles = print_exp_quantiles, x_lab_rotate = x_lab_rotate, plot_padding = plot_padding, flip = flip, k = k, feature_km_repeats = feature_km_repeats, ident_km_repeats = ident_km_repeats, row_label_size = row_label_size, row_label_fontface = row_label_fontface, grid_color = grid_color, cluster_feature = cluster_feature, cluster_ident = cluster_ident, column_label_size = column_label_size, legend_label_size = legend_label_size, legend_title_size = legend_title_size, raster = raster, plot_km_elbow = plot_km_elbow, elbow_kmax = elbow_kmax, assay = assay, group.by = group.by, idents = idents, show_parent_dend_line = show_parent_dend_line, seed = seed) 1: Clustered_DotPlot(seurat_object = Seurat, split.by = "time2", features = c("IL4R", "IL9R", "IL21R", "IL7R"), colors_use_exp = PurpleAndYellow())

table(Seurat@meta.data$time2)
3mo 6mo GMP Wk1 Wk2 Wk3 Wk4 Wk6 Wk8
368 7 118749 21085 17612 19872 5562 1 1535

Code:
Clustered_DotPlot(Seurat,split.by = "time2",features = c("IL4R","IL9R","IL21R","IL7R"),colors_use_exp = PurpleAndYellow())

Thank you!

[samuel-marsh]

Hi @apal6,

Sorry for delay getting back to you. Before we continue trying to troubleshoot back and forth I wanted to check if it is possible to share the object so I can try and troubleshoot directly? If so please send me an email: samuel.marsh@childrens.harvard.edu and I will send you google drive link to upload the object.

If not let me know here and there are couple of things we can try back and forth here.

Best,
Sam

[apal6]

Hi @samuel-marsh ,

I think I got it working. Thank you for your time.

[zhang-HZAU]

hi~I also encountered the same error when using the same code. How did you debug it?

[ishwarvh]

Hello @samuel-marsh ,

Unfortunately, I have come across this same issue in scCustomize_3.0.1.

Clustered_DotPlot( s_obj, features = c( "Pclaf","Nr2f1","Cdk1", "Tbx1","Oc90","Kcnq3", "Ptprt", "Gfra1" ,"E130114P18Rik", "Clu", "Rorb", "Thrb", "Tgfb2", "Adamtsl1", "Thsd7a", "Sema3e", "Fut9", "Nkain2" ), split.by = "orig.ident" )

results in

Error in `[.data.frame`(percent_mat, , idx) : undefined columns selected

Happy to share my seurat object if you can send me a upload link.

Here is my sessionInfo:

sessionInfo

R version 4.4.1 (2024-06-14) Platform: x86_64-pc-linux-gnu Running under: Ubuntu 22.04.5 LTS

Matrix products: default
BLAS: /usr/lib/x86_64-linux-gnu/blas/libblas.so.3.10.0
LAPACK: /usr/lib/x86_64-linux-gnu/lapack/liblapack.so.3.10.0

locale:
[1] LC_CTYPE=en_US.UTF-8 LC_NUMERIC=C LC_TIME=en_US.UTF-8 LC_COLLATE=en_US.UTF-8 LC_MONETARY=en_US.UTF-8
[6] LC_MESSAGES=en_US.UTF-8 LC_PAPER=en_US.UTF-8 LC_NAME=C LC_ADDRESS=C LC_TELEPHONE=C
[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C

time zone: America/Los_Angeles
tzcode source: system (glibc)

attached base packages:
[1] stats4 stats graphics grDevices datasets utils methods base

other attached packages:
[1] SingleCellExperiment_1.26.0 SummarizedExperiment_1.34.0 Biobase_2.64.0 GenomicRanges_1.56.2 GenomeInfoDb_1.40.1
[6] IRanges_2.38.1 S4Vectors_0.42.1 BiocGenerics_0.50.0 MatrixGenerics_1.16.0 matrixStats_1.4.1
[11] EnhancedVolcano_1.22.0 ggrepel_0.9.6 SeuratWrappers_0.4.0 stringr_1.5.1 ggtree_3.12.0
[16] presto_1.0.0 data.table_1.16.4 Rcpp_1.0.13-1 scCustomize_3.0.1 ggplot2_3.5.1
[21] dplyr_1.1.4 Seurat_5.1.0 SeuratObject_5.0.2 sp_2.1-4

loaded via a namespace (and not attached):
[1] RcppAnnoy_0.0.22 splines_4.4.1 later_1.4.1 prismatic_1.1.2 ggplotify_0.1.2 tibble_3.2.1
[7] R.oo_1.27.0 polyclip_1.10-7 janitor_2.2.1 fastDummies_1.7.4 lifecycle_1.0.4 doParallel_1.0.17
[13] globals_0.16.3 lattice_0.22-6 MASS_7.3-64 magrittr_2.0.3 plotly_4.10.4 yaml_2.3.10
[19] remotes_2.5.0 httpuv_1.6.15 sctransform_0.4.1 spam_2.11-0 spatstat.sparse_3.1-0 reticulate_1.40.0
[25] cowplot_1.1.3 pbapply_1.7-2 RColorBrewer_1.1-3 lubridate_1.9.4 abind_1.4-8 zlibbioc_1.50.0
[31] Rtsne_0.17 purrr_1.0.2 R.utils_2.12.3 yulab.utils_0.1.8 circlize_0.4.16 GenomeInfoDbData_1.2.12
[37] irlba_2.3.5.1 listenv_0.9.1 spatstat.utils_3.1-1 tidytree_0.4.6 goftest_1.2-3 RSpectra_0.16-2
[43] spatstat.random_3.3-2 fitdistrplus_1.2-1 parallelly_1.41.0 DelayedArray_0.30.1 leiden_0.4.3.1 codetools_0.2-20
[49] tidyselect_1.2.1 shape_1.4.6.1 aplot_0.2.4 UCSC.utils_1.0.0 farver_2.1.2 spatstat.explore_3.3-3
[55] jsonlite_1.8.9 GetoptLong_1.0.5 progressr_0.15.1 iterators_1.0.14 ggridges_0.5.6 survival_3.8-3
[61] systemfonts_1.1.0 foreach_1.5.2 tools_4.4.1 ragg_1.3.3 treeio_1.28.0 ica_1.0-3
[67] glue_1.8.0 SparseArray_1.4.8 gridExtra_2.3 withr_3.0.2 BiocManager_1.30.25 fastmap_1.2.0
[73] digest_0.6.37 rsvd_1.0.5 timechange_0.3.0 R6_2.5.1 mime_0.12 gridGraphics_0.5-1
[79] textshaping_0.4.1 ggprism_1.0.5 colorspace_2.1-1 Cairo_1.6-2 scattermore_1.2 tensor_1.5
[85] spatstat.data_3.1-4 R.methodsS3_1.8.2 tidyr_1.3.1 generics_0.1.3 renv_1.0.11 S4Arrays_1.4.1
[91] httr_1.4.7 htmlwidgets_1.6.4 uwot_0.2.2 pkgconfig_2.0.3 gtable_0.3.6 ComplexHeatmap_2.20.0
[97] lmtest_0.9-40 XVector_0.44.0 htmltools_0.5.8.1 dotCall64_1.2 clue_0.3-66 scales_1.3.0
[103] png_0.1-8 spatstat.univar_3.1-1 snakecase_0.11.1 ggfun_0.1.8 rstudioapi_0.17.1 rjson_0.2.23
[109] reshape2_1.4.4 nlme_3.1-166 zoo_1.8-12 GlobalOptions_0.1.2 KernSmooth_2.23-26 parallel_4.4.1
[115] miniUI_0.1.1.1 vipor_0.4.7 ggrastr_1.0.2 pillar_1.10.0 grid_4.4.1 vctrs_0.6.5
[121] RANN_2.6.2 promises_1.3.2 xtable_1.8-4 cluster_2.1.8 beeswarm_0.4.0 paletteer_1.6.0
[127] cli_3.6.3 compiler_4.4.1 crayon_1.5.3 rlang_1.1.4 future.apply_1.11.3 labeling_0.4.3
[133] rematch2_2.1.2 plyr_1.8.9 forcats_1.0.0 fs_1.6.5 ggbeeswarm_0.7.2 stringi_1.8.4
[139] viridisLite_0.4.2 deldir_2.0-4 munsell_0.5.1 lazyeval_0.2.2 spatstat.geom_3.3-4 Matrix_1.7-1
[145] RcppHNSW_0.6.0 patchwork_1.3.0 future_1.34.0 shiny_1.10.0 ROCR_1.0-11 igraph_2.1.2
[151] ape_5.8-1

Thank you,
Ishwar

[lengfei5]

same error here with scCustomize_3.0.1

[samuel-marsh]

@lengfei5 @ishwarvh,

Thanks for reports and I’ve reopened the issue. Could you both confirm that the genes you are plotting are expressed by at least one cell in each of the groups you are splitting the plot by?

Best,
Sam

[samuel-marsh]

You can run:

Feature_Present(data = OBJ, features = c("GENE1", "GENE2", ...)

Best,
Sam