Closed Aavdili closed 4 months ago
samuel-marsh
commented
4 months ago Hi,
Not member of dev team but hopefully can be helpful. Can you provide link to public data you are using or reproducible example with matrix extracted from SeuratData object? Could also provide the output of sessionInfo()?
Thanks, Sam
Aavdili
commented
4 months ago Hi the link: https://www.ebi.ac.uk/biostudies/arrayexpress/studies/E-MTAB-11673
and used the: F158_YS_barcodes.tsv.gz, F158_YS_features.tsv.gz, F158_YS_matrix.mtx.gz, and also changed the names of the files -> without (F158_YS)
I managed to create the SeuratObject, and no error is appearing.
and when I check the meta data
view(obj@meta.data), I have just a row and one column with the orig.ident.
[1] "MIR1302-2HG" "AL627309.1" "AL627309.5" "AP006222.2" "LINC01409" "FAM87B" "LINC01128" "LINC00115"
[9] "FAM41C" "AL645608.2" "LINC02593" "SAMD11" "NOC2L" "KLHL17" "PLEKHN1" "AL645608.7"
[17] "HES4" "ISG15" "AL645608.1" "AGRN" "AL645608.8" "RNF223" "C1orf159" "AL390719.3"
[25] "AL390719.2" "TTLL10" "TNFRSF18" "TNFRSF4" "SDF4" "B3GALT6" "C1QTNF12" "AL162741.1"
[33] "UBE2J2" "LINC01786" "SCNN1D" "ACAP3" "PUSL1" "INTS11" "CPTP" "TAS1R3"
[41] "DVL1" "MXRA8" "AURKAIP1" "CCNL2" "MRPL20-AS1" "MRPL20" "AL391244.2" "ANKRD65"
[49] "AL391244.1" "TMEM88B" "LINC01770" "VWA1" "ATAD3C" "ATAD3B" "ATAD3A" "TMEM240"
[57] "SSU72" "AL645728.1" "FNDC10" "AL691432.4" "AL691432.2" "MIB2" "MMP23B" "CDK11B"
[65] "FO704657.1" "SLC35E2B" "CDK11A" "SLC35E2A" "NADK" "GNB1" "AL109917.1" "CALML6"
colnames(obj[])
[1] "AAACCTGAGAAACCGC-WS_wEMB12142156" "AAACCTGAGAGTCTGG-WS_wEMB12142156" "AAACCTGAGATATGCA-WS_wEMB12142156" [4] "AAACCTGAGCAAATCA-WS_wEMB12142156" "AAACCTGAGCGCTCCA-WS_wEMB12142156" "AAACCTGAGCGTCAAG-WS_wEMB12142156" [7] "AAACCTGAGCTGAAAT-WS_wEMB12142156" "AAACCTGAGTACTTGC-WS_wEMB12142156" "AAACCTGAGTAGATGT-WS_wEMB12142156" [10] "AAACCTGAGTGACTCT-WS_wEMB12142156" "AAACCTGAGTGGGATC-WS_wEMB12142156" "AAACCTGCAAGTTCTG-WS_wEMB12142156" [13] "AAACCTGCAATGTAAG-WS_wEMB12142156" "AAACCTGCACATGGGA-WS_wEMB12142156" "AAACCTGCACCACCAG-WS_wEMB12142156" [16] "AAACCTGCACTTCTGC-WS_wEMB12142156" "AAACCTGCAGATCCAT-WS_wEMB12142156" "AAACCTGCAGCTCGCA-WS_wEMB12142156" [19] "AAACCTGCATGCCTAA-WS_wEMB12142156" "AAACCTGCATTCCTGC-WS_wEMB12142156" "AAACCTGGTAAATGTG-WS_wEMB12142156" [22] "AAACCTGGTACGAAAT-WS_wEMB12142156" "AAACCTGGTAGAGGAA-WS_wEMB12142156" "AAACCTGGTAGCTCCG-WS_wEMB12142156"
samuel-marsh
commented
4 months ago Hi,
Thanks for the link. So I'm unable to replicate your issue either with or without renaming the files:
library(tidyverse)
library(Seurat)
library(scCustomize)
> list.files("~/Downloads/E-MTAB-11673/")
[1] "F158_YS_barcodes.tsv.gz" "F158_YS_features.tsv.gz" "F158_YS_matrix.mtx.gz"
> test <- Read10X_GEO("~/Downloads/E-MTAB-11673/")
Reading 10X files from directory
|==================================================| 100% elapsed=23s
> list.files("~/Downloads/testing_rename/")
[1] "barcodes.tsv.gz" "features.tsv.gz" "matrix.mtx.gz"
> test2 <- Read10X("~/Downloads/testing_rename/")
> seurat <- CreateSeuratObject(counts = test, min.cells = 3, min.features = 200)
Warning: Data is of class dgTMatrix. Coercing to dgCMatrix.
> seurat2 <- CreateSeuratObject(counts = test2, min.cells = 3, min.features = 200)
> head(seurat@meta.data)
orig.ident nCount_RNA nFeature_RNA
AAACCTGAGAAACCGC-WS_wEMB12142156 SeuratProject 1647 777
AAACCTGAGAGTCTGG-WS_wEMB12142156 SeuratProject 1246 490
AAACCTGAGATATGCA-WS_wEMB12142156 SeuratProject 989 557
AAACCTGAGCAAATCA-WS_wEMB12142156 SeuratProject 993 362
AAACCTGAGCGCTCCA-WS_wEMB12142156 SeuratProject 1799 1106
AAACCTGAGCGTCAAG-WS_wEMB12142156 SeuratProject 2120 1295
> head(seurat2@meta.data)
orig.ident nCount_RNA nFeature_RNA
AAACCTGAGAAACCGC-WS_wEMB12142156 SeuratProject 1647 777
AAACCTGAGAGTCTGG-WS_wEMB12142156 SeuratProject 1246 490
AAACCTGAGATATGCA-WS_wEMB12142156 SeuratProject 989 557
AAACCTGAGCAAATCA-WS_wEMB12142156 SeuratProject 993 362
AAACCTGAGCGCTCCA-WS_wEMB12142156 SeuratProject 1799 1106
AAACCTGAGCGTCAAG-WS_wEMB12142156 SeuratProject 2120 1295
I would suggest making sure Seurat and your other packages are fully updated and trying to read data in again.
Best, Sam
Aavdili
commented
4 months ago Thank you for getting back to me so quickly.
I was rechecking the packages, and I think it was due to the tidyverse (1.3.2), I updated it to the (2.0.0) and the problem was solved.
I never thought that because our cluster is maintained from the bioinformatic core facility (according to them the packages are always up to date).
Nevertheless. thank you again
Best
I use publicly available scRNA seq data (folder with 3 required files: barcodes.tsv.gz, features.tsv.gz and matrix.mtx.gz)
Code:
df <- Read10X (data.dir = "../path_to _folder/")
obj <- CreateSeuratObject ( counts= df, project= "x", min.cells =3, min.features=200)
then returns just a SeuratObject not containing nCounts_RNA and nFeature_RNA, Just the barcodes in a row and a column with orig.ident.
str(obj)
Formal class 'Seurat' [package "SeuratObject"] with 13 slots ..@ assays :List of 1 .. ..$ RNA:Formal class 'Assay5' [package "SeuratObject"] with 8 slots .. .. .. ..@ layers :List of 1 .. .. .. .. ..$ counts:Formal class 'dgCMatrix' [package "Matrix"] with 6 slots .. .. .. .. .. .. ..@ i : int [1:3155750] 1 13 15 21 24 27 29 33 34 36 ... .. .. .. .. .. .. ..@ p : int [1:1071] 0 4303 7386 10028 12328 14067 18223 21687 24001 26820 ... .. .. .. .. .. .. ..@ Dim : int [1:2] 18057 1070 .. .. .. .. .. .. ..@ Dimnames:List of 2 .. .. .. .. .. .. .. ..$ : NULL .. .. .. .. .. .. .. ..$ : NULL .. .. .. .. .. .. ..@ x : num [1:3155750] 1 3 1 2 1 1 2 1 2 1 ... .. .. .. .. .. .. ..@ factors : list() .. .. .. ..@ cells :Formal class 'LogMap' [package "SeuratObject"] with 1 slot .. .. .. .. .. ..@ .Data: logi [1:1070, 1] TRUE TRUE TRUE TRUE TRUE TRUE ... .. .. .. .. .. .. ..- attr(, "dimnames")=List of 2 .. .. .. .. .. .. .. ..$ : chr [1:1070] "AAACAAGTATCTCCCA-1" "AAACAGAGCGACTCCT-1" "AAACATTTCCCGGATT-1" "AAACGGGCGTACGGGT-1" ... .. .. .. .. .. .. .. ..$ : chr "counts" .. .. .. .. .. ..$ dim : int [1:2] 1070 1 .. .. .. .. .. ..$ dimnames:List of 2 .. .. .. .. .. .. ..$ : chr [1:1070] "AAACAAGTATCTCCCA-1" "AAACAGAGCGACTCCT-1" "AAACATTTCCCGGATT-1" "AAACGGGCGTACGGGT-1" ... .. .. .. .. .. .. ..$ : chr "counts" .. .. .. ..@ features :Formal class 'LogMap' [package "SeuratObject"] with 1 slot .. .. .. .. .. ..@ .Data: logi [1:18057, 1] TRUE TRUE TRUE TRUE TRUE TRUE ... .. .. .. .. .. .. ..- attr(, "dimnames")=List of 2 .. .. .. .. .. .. .. ..$ : chr [1:18057] "RP11-34P13.7" "FO538757.2" "AP006222.2" "RP11-206L10.9" ... .. .. .. .. .. .. .. ..$ : chr "counts" .. .. .. .. .. ..$ dim : int [1:2] 18057 1 .. .. .. .. .. ..$ dimnames:List of 2 .. .. .. .. .. .. ..$ : chr [1:18057] "RP11-34P13.7" "FO538757.2" "AP006222.2" "RP11-206L10.9" ... .. .. .. .. .. .. ..$ : chr "counts" .. .. .. ..@ default : int 1 .. .. .. ..@ assay.orig: chr(0) .. .. .. ..@ meta.data :'data.frame': 18057 obs. of 0 variables .. .. .. ..@ misc :List of 1 .. .. .. .. ..$ calcN: logi FALSE .. .. .. ..@ key : chr "rna_" ..@ meta.data :'data.frame': 1070 obs. of 1 variable: .. ..$ orig.ident: Factor w/ 1 level "yolk_sac": 1 1 1 1 1 1 1 1 1 1 ... ..@ active.assay: chr "RNA" ..@ active.ident: Factor w/ 1 level "yolk_sac": 1 1 1 1 1 1 1 1 1 1 ... .. ..- attr(*, "names")= chr [1:1070] "AAACAAGTATCTCCCA-1" "AAACAGAGCGACTCCT-1" "AAACATTTCCCGGATT-1" "AAACGGGCGTACGGGT-1" ... ..@ graphs : list() ..@ neighbors : list() ..@ reductions : list() ..@ images : list() ..@ project.name: chr "yolk_sac" ..@ misc : list() ..@ version :Classes 'package_version', 'numeric_version' hidden list of 1 .. ..$ : int [1:3] 5 0 1 ..@ commands : list() ..@ tools : list()