Not getting the reference part of the heatmap

[Cristinacondel1]

Dear All,

I'm running the code for inferCNV and everything gets generated correctly but in all the heatmaps generated the reference cell heatmap is missing.

The code I'm using for the run is as follows:

# Create the inferCNV object
  infercnv_obj <- CreateInfercnvObject(
    raw_counts_matrix = HNSC_matrix,
    annotations_file = annotation_file_path,
    delim = ",",
    gene_order_file = "/omics/odcf/analysis/OE0509_projects/hnscc/single_cell/Single_Cell_Analysis/Single_Cell_Datasets/All_Cells_Together/Integration/InferCNV/hg38_gencode_v27.txt",
    ref_group_names = c("Immune", "Stromal"),
    chr_exclude = c("chrM")
  )

   # Perform inferCNV operations to reveal CNV signal
   
  infercnv_obj <- infercnv::run(
    infercnv_obj,
    cutoff = 0.1,
    out_dir = paste0("output_dir_", dataset_id), 
    cluster_by_groups = TRUE,
    denoise = TRUE,
    HMM = TRUE
  )

At one point I got the following error message, but already all the heatmaps were generated:

Error in heatmap.cnv(obs_data, Rowv = obs_dendrogram, Colv = FALSE, cluster.by.row = TRUE,  :
  'RowIndividualColors' must be a character vector of length nrow(x)
Calls: <Anonymous> ... plot_subclusters -> plot_cnv -> .plot_cnv_observations -> heatmap.cnv
In addition: Warning messages:
1: In cbind(row_groupings, get_group_color_palette()(length(table(hcl_obs_annotations_groups)))[hcl_obs_annotations_groups]) :
  number of rows of result is not a multiple of vector length (arg 2)
2: In cbind(split_groups, row_groupings[, 1], hcl_obs_annotations_groups,  :
  number of rows of result is not a multiple of vector length (arg 3)
Execution halted 

But I'm not sure how to proceed after trying out and checking that all input files are in the right format.

[hayden-bell]

I had the same issue. It is possible that it is related to the size of the your reference and the Cairo back-end is then unable to generate a matrix of that size.

See https://github.com/broadinstitute/infercnv/wiki/Infercnv-runs-successfully-and-generated-plots-but-they-are-blank

[peizhu1205]

output_format="pdf": Changing the output format to PDF can resolve this issue.