A-legac45 commented 10 months ago

Hello I am also troubling with the fragment files which are the output of cellranger 10x multiomic data

seurat_atac <- ArchR2Signac(

ArchRProject = project_Peaks_MACS2_RES0.9,
refversion = "mm10",
#samples = samplelist, # list of samples in the ArchRProject (default will use ArchRProject@cellColData$Sample but another list can be provided)
fragments_dir = inputFiles,
pm = pkm, # peak matrix from getPeakMatrix(),
fragments_file_extension = '_fragments.tsv.gz',
fragments_fromcellranger = "YES", # fragments_fromcellranger This is an Yes or No selection ("NO" | "N" | "No" or "YES" | "Y" | "Yes")
annotation = annotation # annotation from getAnnotation()
)
[1] "In Progress:"
[1] "Prepare Seurat list for each sample"
[1] "First_try_multiomic_archr"
[1] 121983 3733
Error in CreateFragmentObject(path = fragments, cells = cells, validate.fragments = validate.fragments, :
Fragment file does not exist.

For InputFiles I try many things InputFiles <- c("/Users/alegac/Library/CloudStorage/OneDrive-INSTITUTCURIE/Projet_linda_mutiomic_novembre/KDI_2017426_2023-11-15_16-59-39/") InputFiles <- c("/Users/alegac/Library/CloudStorage/OneDrive-INSTITUTCURIE/Projet_linda_mutiomic_novembre/KDI_2017426_2023-11-15_16-59-39") InputFiles <- c("/Users/alegac/Library/CloudStorage/OneDrive-INSTITUTCURIE/Projet_linda_mutiomic_novembre/KDI_2017426_2023-11-15_16-59-39/atac_fragments.tsv.gz")

Thanks for help

rootze commented 10 months ago

Hello @A-legac45, Thanks for using ArchRtoSignac package

Before everything, do you only have one sample? If so, please update your ArchRtoSignac package; I recently encountered some requests with working on only one sample, so made a minor update in the ArchRtoSignac package, hope it works. BE CAUTIOUS when you update the ArchRtoSignac package, please don't update anything else; recent updates of the Seurat and SeuratObject packages may change something for other packages supporting Seurat V4.

Maybe try this from ****STEP 3 Option2:

fragments_dirs <- c(
"/Users/alegac/Library/CloudStorage/OneDrive-INSTITUTCURIE/Projet_linda_mutiomic_novembre/KDI_2017426_2023-11-15_16-59-39/atac_"
)

# Call the ArchR2Signac function with the provided arguments
seurat_atac <- ArchR2Signac(
  ArchRProject = project_Peaks_MACS2_RES0.9,
  refversion = "mm10",
  fragments_dir = fragments_dirs,
  pm = pkm,
  fragments_fromcellranger = "NO",
  fragments_file_extension = 'fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz
  annotation = annotations
)

I see the fragments_fromcellranger parameter is a bit confusing when choosing NO, fragments_fromcellranger = 'NO' is for fragments from NON Cellranger ATAC output or even from Cellranger ATAC output but don't match the standard output PATH from Cellranger ATAC cellranger-atac count

Please let me know if it works or not, more than happy to help you more.

A-legac45 commented 10 months ago

Hello I have only one sample. Multiomic 10x dataset

I try this but still not working :

fragments_dirs <- c(

"/Users/alegac/Documents/archr_linda_multiom_dec_2023/"

)

or

fragments_dirs <- c( "/Users/alegac/Documents/archr_linda_multiom_dec2023/atac" )

seurat_atac <- ArchR2Signac(

ArchRProject = project_Peaks_MACS2_RES0.5,

refversion = "mm10",

fragments_dir = fragments_dirs,

pm = pkm,

fragments_fromcellranger = "YES",

fragments_file_extension = 'atac_fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz

annotation = annotation

) [1] "In Progress:" [1] "Prepare Seurat list for each sample" [1] "First_try_multiomic_archr" [1] 121983 3733 Error in CreateFragmentObject(path = fragments, cells = cells, validate.fragments = validate.fragments, : Fragment file does not exist.

A-legac45 commented 10 months ago

Hello I have only one sample. Multiomic 10x dataset

I try this but still not working :

fragments_dirs <- c(

"/Users/alegac/Documents/archr_linda_multiom_dec_2023/"
)

or

fragments_dirs <- c( "/Users/alegac/Documents/archr_linda_multiom_dec2023/atac" )

seurat_atac <- ArchR2Signac(

ArchRProject = project_Peaks_MACS2_RES0.5,
refversion = "mm10",
fragments_dir = fragments_dirs,
pm = pkm,
fragments_fromcellranger = "YES",
fragments_file_extension = 'atac_fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz
annotation = annotation
) [1] "In Progress:" [1] "Prepare Seurat list for each sample" [1] "First_try_multiomic_archr" [1] 121983 3733 Error in CreateFragmentObject(path = fragments, cells = cells, validate.fragments = validate.fragments, : Fragment file does not exist.

here you will find the structure of my folder "/Users/alegac/Documents/archr_linda_multiom_dec_2023/

De : Ze @.> Envoyé : jeudi 21 décembre 2023 21:41 À : swaruplabUCI/ArchRtoSignac @.> Cc : Le Gac Anne-Laure @.>; Mention @.> Objet : Re: [swaruplabUCI/ArchRtoSignac] Frangments dir with version 1.0.3 (Issue #33)

Hello @A-legac45https://github.com/A-legac45, Thanks for using ArchRtoSignac package

Before everything, do you only have one sample? If so, please update your ArchRtoSignac package; I recently encountered some requests with working on only one sample, so made a minor update in the ArchRtoSignac package, hope it works. BE CAUTIOUS when you update the ArchRtoSignac package, please don't update anything else; recent updates of the Seurat and SeuratObject packages may change something for other packages supporting Seurat V4.

Maybe try this from ****STEP 3 Option2:

fragments_dirs <- c( "/Users/alegac/Library/CloudStorage/OneDrive-INSTITUTCURIE/Projet_linda_mutiomic_novembre/KDI_2017426_2023-11-1516-59-39/atac" )

Call the ArchR2Signac function with the provided arguments

seurat_atac <- ArchR2Signac( ArchRProject = project_Peaks_MACS2_RES0.9, refversion = "mm10", fragments_dir = fragments_dirs, pm = pkm, fragments_fromcellranger = "NO", fragments_file_extension = 'fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz annotation = annotations )

I see the fragments_fromcellranger parameter is a bit confusing when choosing NO, fragments_fromcellranger = 'NO' is for fragments from NON Cellranger ATAC output or even from Cellranger ATAC output but don't match the standard output PATH from Cellranger ATAC cellranger-atac count

Please let me know if it works or not, more than happy to help you more.

— Reply to this email directly, view it on GitHubhttps://github.com/swaruplabUCI/ArchRtoSignac/issues/33#issuecomment-1866903876, or unsubscribehttps://github.com/notifications/unsubscribe-auth/AYDTLNX4MYSIEOWCAQBFKDTYKSNGVAVCNFSM6AAAAABA6LDDH2VHI2DSMVQWIX3LMV43OSLTON2WKQ3PNVWWK3TUHMYTQNRWHEYDGOBXGY. You are receiving this because you were mentioned.Message ID: @.***>

A-legac45 commented 10 months ago

fragments <- CreateFragmentObject(path = fragments_dirs, cells = Cells(pkm), validate.fragments = TRUE) Computing hash Checking for 3733 cell barcodes Error in CreateFragmentObject(path = fragments_dirs, cells = Cells(pkm), : Not all cells requested could be found in the fragment file. fragments <- CreateFragmentObject(path = fragments_dirs, cells = Cells(pkm), validate.fragments = F) Computing hash all(colnames(pkm) %in% Cells(fragments)) [1] TRUE

De : Le Gac Anne-Laure @.> Envoyé : jeudi 21 décembre 2023 22:54 À : swaruplabUCI/ArchRtoSignac @.> Objet : RE: [swaruplabUCI/ArchRtoSignac] Frangments dir with version 1.0.3 (Issue #33)

Hello I have only one sample. Multiomic 10x dataset

I try this but still not working :

fragments_dirs <- c(

"/Users/alegac/Documents/archr_linda_multiom_dec_2023/"
)

or

fragments_dirs <- c( "/Users/alegac/Documents/archr_linda_multiom_dec2023/atac" )

seurat_atac <- ArchR2Signac(

ArchRProject = project_Peaks_MACS2_RES0.5,
refversion = "mm10",
fragments_dir = fragments_dirs,
pm = pkm,
fragments_fromcellranger = "YES",
fragments_file_extension = 'atac_fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz
annotation = annotation
) [1] "In Progress:" [1] "Prepare Seurat list for each sample" [1] "First_try_multiomic_archr" [1] 121983 3733 Error in CreateFragmentObject(path = fragments, cells = cells, validate.fragments = validate.fragments, : Fragment file does not exist.

here you will find the structure of my folder "/Users/alegac/Documents/archr_linda_multiom_dec_2023/

De : Ze @.> Envoyé : jeudi 21 décembre 2023 21:41 À : swaruplabUCI/ArchRtoSignac @.> Cc : Le Gac Anne-Laure @.>; Mention @.> Objet : Re: [swaruplabUCI/ArchRtoSignac] Frangments dir with version 1.0.3 (Issue #33)

Hello @A-legac45https://github.com/A-legac45, Thanks for using ArchRtoSignac package

Before everything, do you only have one sample? If so, please update your ArchRtoSignac package; I recently encountered some requests with working on only one sample, so made a minor update in the ArchRtoSignac package, hope it works. BE CAUTIOUS when you update the ArchRtoSignac package, please don't update anything else; recent updates of the Seurat and SeuratObject packages may change something for other packages supporting Seurat V4.

Maybe try this from ****STEP 3 Option2:

fragments_dirs <- c( "/Users/alegac/Library/CloudStorage/OneDrive-INSTITUTCURIE/Projet_linda_mutiomic_novembre/KDI_2017426_2023-11-1516-59-39/atac" )

Call the ArchR2Signac function with the provided arguments

seurat_atac <- ArchR2Signac( ArchRProject = project_Peaks_MACS2_RES0.9, refversion = "mm10", fragments_dir = fragments_dirs, pm = pkm, fragments_fromcellranger = "NO", fragments_file_extension = 'fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz annotation = annotations )

I see the fragments_fromcellranger parameter is a bit confusing when choosing NO, fragments_fromcellranger = 'NO' is for fragments from NON Cellranger ATAC output or even from Cellranger ATAC output but don't match the standard output PATH from Cellranger ATAC cellranger-atac count

Please let me know if it works or not, more than happy to help you more.

— Reply to this email directly, view it on GitHubhttps://github.com/swaruplabUCI/ArchRtoSignac/issues/33#issuecomment-1866903876, or unsubscribehttps://github.com/notifications/unsubscribe-auth/AYDTLNX4MYSIEOWCAQBFKDTYKSNGVAVCNFSM6AAAAABA6LDDH2VHI2DSMVQWIX3LMV43OSLTON2WKQ3PNVWWK3TUHMYTQNRWHEYDGOBXGY. You are receiving this because you were mentioned.Message ID: @.***>

rootze commented 10 months ago

Hello @A-legac45,

I cannot see your attached screenshots. I don't think GitHub supports that with email replies. However, please double-check what I provided for you in my last reply. I made changes not only to fragments_dirs but also fragments_fromcellranger and fragments_file_extension

# Once again here is the code that I changed for you earlier 
fragments_dirs <- c(
"/Users/alegac/Library/CloudStorage/OneDrive-INSTITUTCURIE/Projet_linda_mutiomic_novembre/KDI_2017426_2023-11-15_16-59-39/atac_"
)

# Call the ArchR2Signac function with the provided arguments
seurat_atac <- ArchR2Signac(
  ArchRProject = project_Peaks_MACS2_RES0.9,
  refversion = "mm10",
  fragments_dir = fragments_dirs,
  pm = pkm,
  fragments_fromcellranger = "NO",
  fragments_file_extension = 'fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz
  annotation = annotations
)

BUT What you had is

seurat_atac <- ArchR2Signac(
      ArchRProject = project_Peaks_MACS2_RES0.5,
       refversion = "mm10",
       fragments_dir = fragments_dirs,
       pm = pkm,
  *   fragments_fromcellranger = "YES",
  *   fragments_file_extension = 'atac_fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz
       annotation = annotation
      )

A-legac45 commented 10 months ago

It still not working

fragments_dirs <- c(

"/Users/alegac/Documents/archr_linda_multiom_dec2023/atac"

) seurat_atac <- ArchR2Signac(

ArchRProject = project_Peaks_MACS2_RES0.5,

refversion = "mm10",

fragments_dir = fragments_dirs,

pm = pkm,

fragments_fromcellranger = "NO",

fragments_file_extension = 'fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz

annotation = annotation

) [1] "IF selecting NO, please make sure to provide fragments_file_extension" [1] "In Progress:" [1] "Prepare Seurat list for each sample" [1] "First_try_multiomic_archr" [1] 121983 3733 Error in CreateFragmentObject(path = fragments, cells = cells, validate.fragments = validate.fragments, : Fragment file does not exist.

Do you want me too send you my archr project or my fragment files??

De : Ze @.> Envoyé : jeudi 21 décembre 2023 23:08 À : swaruplabUCI/ArchRtoSignac @.> Cc : Le Gac Anne-Laure @.>; Mention @.> Objet : Re: [swaruplabUCI/ArchRtoSignac] Frangments dir with version 1.0.3 (Issue #33)

Hello @A-legac45https://github.com/A-legac45,

I cannot see your attached screenshots. I don't think GitHub supports that with email replies. However, please double-check what I provided for you in my last reply. I made changed not only to fragments_dirs but also fragments_fromcellranger and fragments_file_extension

Once again here is the code that I changed for you earlier

fragments_dirs <- c( "/Users/alegac/Library/CloudStorage/OneDrive-INSTITUTCURIE/Projet_linda_mutiomic_novembre/KDI_2017426_2023-11-1516-59-39/atac" )

Call the ArchR2Signac function with the provided arguments

seurat_atac <- ArchR2Signac( ArchRProject = project_Peaks_MACS2_RES0.9, refversion = "mm10", fragments_dir = fragments_dirs, pm = pkm, fragments_fromcellranger = "NO", fragments_file_extension = 'fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz annotation = annotations )

BUT What you had is

seurat_atac <- ArchR2Signac( ArchRProject = project_Peaks_MACS2_RES0.5, refversion = "mm10", fragments_dir = fragments_dirs, pm = pkm,

fragments_fromcellranger = "YES",
fragments_file_extension = 'atac_fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz annotation = annotation )

— Reply to this email directly, view it on GitHubhttps://github.com/swaruplabUCI/ArchRtoSignac/issues/33#issuecomment-1866991802, or unsubscribehttps://github.com/notifications/unsubscribe-auth/AYDTLNU37RQDSDUEVGSS5J3YKSXPRAVCNFSM6AAAAABA6LDDH2VHI2DSMVQWIX3LMV43OSLTON2WKQ3PNVWWK3TUHMYTQNRWHE4TCOBQGI. You are receiving this because you were mentioned.Message ID: @.***>

rootze commented 10 months ago

fragments_dirs <- c(

"/Users/alegac/Documents/archr_linda_multiom_dec2023/atac" + )

@A-legac45 It seems to me that your fragment PATH has changed. Can you show me what your PATH to the files and what files are inside that folder?

A-legac45 commented 10 months ago

I try this :

fragments_dirs <- c("/Users/alegac/Documents/archr_linda_multiom_dec2023/atac")

seurat_atac <- ArchR2Signac( ArchRProject = project_Peaks_MACS2_RES0.5, refversion = "BSgenome.Mmusculus.UCSC.mm10", fragments_dir = fragments_dirs, pm = pkm, fragments_fromcellranger = "NO", fragments_file_extension = 'fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz annotation = annotation )

[1] "IF selecting NO, please make sure to provide fragments_file_extension" [1] "In Progress:" [1] "Prepare Seurat list for each sample" [1] "First_try_multiomic_archr" [1] 121983 3733 Error in CreateFragmentObject(path = fragments, cells = cells, validate.fragments = validate.fragments, : Fragment file does not exist.

the folder contain the following files [cid:a1ac8f48-3f18-4a3a-9167-039b6bd8f9b2]

De : Ze @.> Envoyé : vendredi 22 décembre 2023 00:26 À : swaruplabUCI/ArchRtoSignac @.> Cc : Le Gac Anne-Laure @.>; Mention @.> Objet : Re: [swaruplabUCI/ArchRtoSignac] Frangments dir with version 1.0.3 (Issue #33)

fragments_dirs <- c(

"/Users/alegac/Documents/archr_linda_multiom_dec2023/atac" + )

@A-legac45https://github.com/A-legac45 It seems to me that your fragment PATH has changed. Can you show me what your PATH to the files and what files are inside that folder?

— Reply to this email directly, view it on GitHubhttps://github.com/swaruplabUCI/ArchRtoSignac/issues/33#issuecomment-1867047655, or unsubscribehttps://github.com/notifications/unsubscribe-auth/AYDTLNRXENRH26L3ZHLJULDYKTATLAVCNFSM6AAAAABA6LDDH2VHI2DSMVQWIX3LMV43OSLTON2WKQ3PNVWWK3TUHMYTQNRXGA2DONRVGU. You are receiving this because you were mentioned.Message ID: @.***>

A-legac45 commented 10 months ago

Hello Ze,

I have tried this also I do not know if this can help :

fragments_dirs <- c("/Users/alegac/Documents/archr_linda_multiom_dec_2023/atac_fragments.tsv.gz") fragments <- CreateFragmentObject(path = fragments_dirs, cells = Cells(pkm), validate.fragments = F) Computing hash

all(colnames(pkm) %in% Cells(fragments)) [1] TRUE

fragments <- CreateFragmentObject(path = fragments_dirs, cells = Cells(pkm), validate.fragments = T) Computing hash Checking for 3733 cell barcodes Error in CreateFragmentObject(path = fragments_dirs, cells = Cells(pkm), : Not all cells requested could be found in the fragment file.

The achr projet is filtered may be this is were the problem come from?

De : Le Gac Anne-Laure @.> Envoyé : vendredi 22 décembre 2023 10:26 À : swaruplabUCI/ArchRtoSignac @.> Objet : RE: [swaruplabUCI/ArchRtoSignac] Frangments dir with version 1.0.3 (Issue #33)

I try this :

fragments_dirs <- c("/Users/alegac/Documents/archr_linda_multiom_dec2023/atac")

seurat_atac <- ArchR2Signac( ArchRProject = project_Peaks_MACS2_RES0.5, refversion = "BSgenome.Mmusculus.UCSC.mm10", fragments_dir = fragments_dirs, pm = pkm, fragments_fromcellranger = "NO", fragments_file_extension = 'fragments.tsv.gz', # instead of using fragments_file_extension (.tsv.gz or .fragments.tsv.gz), here just use the whole name fragments.tsv.gz annotation = annotation )

[1] "IF selecting NO, please make sure to provide fragments_file_extension" [1] "In Progress:" [1] "Prepare Seurat list for each sample" [1] "First_try_multiomic_archr" [1] 121983 3733 Error in CreateFragmentObject(path = fragments, cells = cells, validate.fragments = validate.fragments, : Fragment file does not exist.

the folder contain the following files [cid:a1ac8f48-3f18-4a3a-9167-039b6bd8f9b2]

De : Ze @.> Envoyé : vendredi 22 décembre 2023 00:26 À : swaruplabUCI/ArchRtoSignac @.> Cc : Le Gac Anne-Laure @.>; Mention @.> Objet : Re: [swaruplabUCI/ArchRtoSignac] Frangments dir with version 1.0.3 (Issue #33)

fragments_dirs <- c(

"/Users/alegac/Documents/archr_linda_multiom_dec2023/atac" + )

@A-legac45https://github.com/A-legac45 It seems to me that your fragment PATH has changed. Can you show me what your PATH to the files and what files are inside that folder?

— Reply to this email directly, view it on GitHubhttps://github.com/swaruplabUCI/ArchRtoSignac/issues/33#issuecomment-1867047655, or unsubscribehttps://github.com/notifications/unsubscribe-auth/AYDTLNRXENRH26L3ZHLJULDYKTATLAVCNFSM6AAAAABA6LDDH2VHI2DSMVQWIX3LMV43OSLTON2WKQ3PNVWWK3TUHMYTQNRXGA2DONRVGU. You are receiving this because you were mentioned.Message ID: @.***>

rootze commented 10 months ago

The achr projet is filtered may be this is were the problem come from?

Hello @A-legac45,

It could be. I am not sure. I have very limited experience with true multi-omic datasets. Are you using a public dataset?

mitchTwoTimes commented 1 month ago

Hi @A-legac45 and @rootze,

If this is still an issue I think I was able to get it to work by going in to the ArchR2Signac function and changing a few things to match the cell ranger ARC output.

Line 12: output_dir = "/croo_output/"

Lines 15/16 :

cur_fragments <- ifelse(is.list(fragments_dir), paste0(fragments_dir[[which(samples == 
                                                                                    cur_sample)]], output_dir, "atac_fragments.tsv.gz"),

swaruplabUCI / ArchRtoSignac

Frangments dir with version 1.0.3 #33

Call the ArchR2Signac function with the provided arguments

Call the ArchR2Signac function with the provided arguments

Once again here is the code that I changed for you earlier

Call the ArchR2Signac function with the provided arguments