tSNE plots from the full run of cluster_map won't open

[lucygarner]

Hi,

I have run cluster_map and it successfully generated the output files.

res <- cluster_map(marker_file_list, edge_cutoff = 0.1, output = "../results/clustermap_full", single_obj_list = single_obj_list, comb_obj = comb_obj, comb_delim = "-")

Whilst I can open the circos plot, hcluster and heatmap, none of the .tsne.pdf files will open. They are all listed as 4 kb in size and when I try to open them, I get the following error message:

Any thoughts as to how to fix this?

Best wishes,
Lucy

[owenkuo]

any error or warning?

[jmrussell]

Hi Lucy,

Can you tell me what version of Seurat you're using?

[lucygarner]

Hi both,

Thank you for the response. There is no error message and I am using Seurat v3.1.3.

Best wishes,
Lucy

[owenkuo]

Hi jmrussell,

I've tested the package, and got some trouble during the stage below

res <- cluster_map(marker_file_list, edge_cutoff = 0.1, output = 'combine', single_obj_list = single_obj_list, comb_obj = objList$combine)

an error occurs as follow,
Error in DimPlot(object = new("Seurat", assays = list(RNA = new("Assay", : unused arguments

and the same as Lucy I got the blank recolor pic of sample01

I used Seurat 3.1.4 to create the .rds of two single obj and a Seurat-merged combine obj.

Thanks for repply

Owen

Hi Lucy,

Can you tell me what version of Seurat you're using?

[jmrussell]

Hi jmrussell,

I've tested the package, and got some trouble during the stage below

res <- cluster_map(marker_file_list, edge_cutoff = 0.1, output = 'combine', single_obj_list = single_obj_list, comb_obj = objList$combine)

an error occurs as follow,
Error in DimPlot(object = new("Seurat", assays = list(RNA = new("Assay", : unused arguments

and the same as Lucy I got the blank recolor pic of sample01

I used Seurat 3.1.4 to create the .rds of two single obj and a Seurat-merged combine obj.

Thanks for repply

Owen

Hi Lucy,
Can you tell me what version of Seurat you're using?

Thanks, I will take a look at this weekend and take care of it.

[jmrussell]

Hi jmrussell,

I've tested the package, and got some trouble during the stage below

res <- cluster_map(marker_file_list, edge_cutoff = 0.1, output = 'combine', single_obj_list = single_obj_list, comb_obj = objList$combine)

an error occurs as follow,
Error in DimPlot(object = new("Seurat", assays = list(RNA = new("Assay", : unused arguments

and the same as Lucy I got the blank recolor pic of sample01

I used Seurat 3.1.4 to create the .rds of two single obj and a Seurat-merged combine obj.

Thanks for repply

Owen

Hi Lucy,
Can you tell me what version of Seurat you're using?

Hi jmrussell,

I've tested the package, and got some trouble during the stage below

res <- cluster_map(marker_file_list, edge_cutoff = 0.1, output = 'combine', single_obj_list = single_obj_list, comb_obj = objList$combine)

an error occurs as follow,
Error in DimPlot(object = new("Seurat", assays = list(RNA = new("Assay", : unused arguments

and the same as Lucy I got the blank recolor pic of sample01

I used Seurat 3.1.4 to create the .rds of two single obj and a Seurat-merged combine obj.

Thanks for repply

Owen

Hi Lucy,
Can you tell me what version of Seurat you're using?

@owenkuo

Did you use the SCTransform normalization or the previous method of NormalizeData/ScaleData/FindVariableFeatures?

[owenkuo]

Hi @jmrussell ,

Yes, I have done NormalizeData/FindVariableFeatures/ScaleData for each sample and combine obj.

Owen

[lucygarner]

Hi @jmrussell,

I used SCTransform for my analysis.

Best,
Lucy

[owenkuo]

Hi @jmrussell

it seems at recolor step the code for Seuratobj v2 the label function at do.label, but at Seurat v3 is just label?

Owen

[jmrussell]

Hi @jmrussell

it seems at recolor step the code for Seuratobj v2 the label function at do.label, but at Seurat v3 is just label?

Owen

Hi Owen,

Can you try again with your dataset and let me know if it works now? Seurat actually deprecated TSNEPlot and replaced it with DimPlot. I think I've put a check for which version of Seurat is running that should fix this.

[owenkuo]

Hi @jmrussell ,

Thanks for the modified of the code, but it seems something wrong while the recolor.r parsing

Error in parse(outFile) :
C:/Users/owen/AppData/Local/Temp/RtmpaePF5T/R.INSTALL59472b86ec6/ClusterMap/R/recolor.R:52:4: unexpected 'else'
51: }
52: else
^
ERROR: unable to collate and parse R files for package 'ClusterMap'

but still thanks for the continue update

[jmrussell]

Thanks Owen, give it another shot now. Unfortunately I am not at my work environment so I made a small error when pushing the new changes. It should install properly now. I also don't have suitable test data for this since I can't access it right now. If you're using a public dataset that you can link me I can test this for you as well.

[owenkuo]

Hi @jmrussell,

Thanks for the update! It seems good for help.
Now, just got a noted error

Error in DimPlot(obj, label = T, label.size = 8, group.by = "regroup", :
unused arguments (colors.use = color[sort(as.numeric(unique(new_group)))], plot.title = toupper(output))

How can I fix this?

Owen

[jmrussell]

It should be fixed now. It turns out there were 3 issues happening - the plots were not saving properly so I changed the way they are being saved which should resolve that issue, and I also cleaned up a few of the new Seurat functions we rely on so that that you shouldn't get errors like that anymore. You might get a few warnings I plan on cleaning up in the future but you can ignore them safely now.

Will you let me know if you're able to run your data through without issue and generate visible plots again?

Thanks for your help.

[lucygarner]

Hi @jmrussell,

Thank you for looking into this. The full run seems to be working for me now and the tSNE plots are generated as expected.

However, I am now getting an error with the simple run.

res <- cluster_map(marker_file_list, edge_cutoff = 0.1, output = "../results/clustermap")

The marker file list is just a character vector containing the paths to the output of FindMarkers for each of seven individual samples.

ERROR:
Error in cluster_map(marker_file_list, edge_cutoff = 0.1, output = "../results/clustermap") :
trying to get slot "version" from an object of a basic class ("NULL") with no slots

Best,
Lucy