Loading 10x data into Monocle 3
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21 months ago
akh22 ▴ 50

Hi,

I've been struggling to load our mouse 10x data into Monocle3 by using;

cds10x<-load_cellranger_data("/Volumes/Aura/monocle/")

However, I get this error message;

Error in get_genome_in_matrix_path(matrix_dir, genome) : 
Multiple genomes found; please specify one. 
Genomes present: barcodes.tsv, features.tsv, matrix.mtx

My directory structure of data looks as follows;

/Volumes/Documents/ahoji/Desktop/Aura 2019-06-28 10-50-28.png

I tried several genome= GRCn38, mm10 and so forth but got the same error. I followed loading 10x PBMC exmple in Dave Tangs website like this,

/Volumes/Documents/ahoji/Desktop/Aura 2019-06-28 10-56-31.png

then I got a following error;

Error in load_cellranger_data("/Volumes/Aura/monocle/") :

Gene name or features file missing

I even created a SingleCellExperiment object and used new_cell_data_set ;

sce<-read10xCounts("/Volumes/Aura/scRNA/monocle/filtered_gene_bc_matrices/")
sce.counts<-sce@assays@data@listData$counts
gene_meta_data<-rowData(sce)
cell_meta_data<-colData(sce)
testcds<-new_cell_data_set(sce.counts,  cell_metadata = cell_metadata, gene_metadata = gene_metadata)

as written in Monocle3 intro web site but I got ;

Error in new_cell_data_set(sce.counts, cell_metadata = cell_metadata,  : 
object 'cell_metadata' not found

It's been a real struggle and painful. I did not have any issues with Seurat3. Anyway, I'd really really appreciate any pointers or suggestions.

RNA-Seq next-gen R Monocle3 10xdata • 2.4k views
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Please read How to add images to a Biostars post to learn how to add images to a post. Copy pasting the desktop file path will not work.

But the path does help in debugging your problem - See how it says /Volumes/Documents/ahoji? I have a feeling Volumes/Aura/ is not a valid path - can you double check that please?

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They are the right paths. /Volumes/Documents/ahoji as a part of "/Volumes/Documents/ahoji/Desktop/Aura 2019-06-28 10-50-28.png" is a path to my image file showing a directory structure within Volumes/Aura/monocle/ but for some reasons the image was not loaded.

Originally, I had the following directory structure :

monocle
└── outs
    └── filtered_gene_bc_matrices
        ├── barcodes.tsv
        ├── features.tsv
        └── matrix.mtx

I modified it to a following after following 10x 8k PBMC example.

monocle
└── outs
    └── filtered_gene_bc_matrices
        └── GRCm38
            ├── barcodes.tsv
            ├── features.tsv
            └── matrix.mtx
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but for some reasons the image was not loaded

It did not load because you did not upload the image. Try following the guide I pointed out to you. That will help us see the images you're trying to share.

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Ok Here we go;

Directory Structure 1

Directory Structure 2

and the location of monocle/ is /Volumes/Aura/monocle

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Their website says the tool is in beta phase. I think emailing the google group mentioned on the site is a good option.

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Thanks. I don't see any posts in their google group, and that's why I posted a question here, but I sent email to monocle-user to see if I can get faster response.

Thanks.

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Entering edit mode
21 months ago
akh22 ▴ 50

I figured it out. It was simple. CellRanger uses mm10 as a mouse ref, genome, and all I have to do is to change the name of folder, GRCm38 to mm10. This did it.

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