Unclear how to specify BAM files in sample sheet

[mschilli87]

Hi, I'm trying to run SCALPEL with subsampled data. At this time, I'm trying to run SCALPEL with BAM file for shorter running time. So I made my samplesheet.csv file like this format: sample, library_info, barcode, bam file. But I got error related parameter recognition problem about my sample sheet. Even though I changed the format into sample, NA, NA, CellRanger_output (contains BAM file and cellRanger output), I have similar problems. So, Do you have any regularized format when user provides BAM file as an input?

Originally posted by @Choeun-fran in #27

@Choeun-fran: I moved this into a new issue since it doesn't appear related in any way to your earlier issue.

Unfortunetly, I don't quite understand what is your problem.
Please provide us with all details requried to reproduce your error on our systems, otherwise we won't be able to help.
In this case, at least we need to know the error message you get, the sample file you use, the command you use to invoke SCALPEL, and which vesion of SCALPEL you are using.
Also, before posting your response, please read the GitHub formatting guide so you can format your comment in a way that makes it easier for us to read and copy/paste from (e.g compare your

[...] I changed the format into sample, NA, NA, CellRanger_output (contains BAM file and cellRanger output) [...]

to

[...] I changed the format into sample, NA, NA, CellRanger_output (contains BAM file and cellRanger output) [...]

).

[Choeun-fran]

Oh, I'm sorry it's my fault.
I tried to run SCALPEL with one of my samples. This is the format of samplesheet.csv I made:

PO23,NA,NA,/node200data/dhkim1674/SC_SR_Samsung_Alzheimer/cellranger_output/PO23/

(sample, NA, NA, cellRanger outputs including BAM and cellRanger matrix)

When I ran SCALPEL with this samplesheet.csv. I got an error message I described below:

ERROR ~ Argument of file function cannot be empty -- Check script '/home/dlchdms0109/Isoform_imputation/sr_tx_analysis/scalpel/SCALPEL/main.nf' at line: 117 or see '.nextflow.log' file for more details All arguments should be occupied.

I also tried with different format of samplesheet.csv like sample, library_id, barcodes_path, bam_path

PO23, PO23, /node200data/dhkim1674/SC_SR_Samsung_Alzheimer/cellranger_output/PO23/outs/filtered_feature_bc_matrix/barcodes.tsv.gz, /node200data/dhkim1674/SC_SR_Samsung_Alzheimer/cellranger_output/PO23/outs/possorted_genome_bam.bam

After second trial, I had another error message:

 Command error: Attaching package: ‘dplyr’ The following objects are masked from ‘package:data.table’: between, first, last The following objects are masked from ‘package:stats’: filter, lag The following objects are masked from ‘package:base’: intersect, setdiff, setequal, union Loading required package: SeuratObject Loading required package: sp Attaching package: ‘SeuratObject’ The following objects are masked from ‘package:base’: intersect, t [1] "reading file..." Error in Read10X(paste0(args$repo_path, "/outs/filtered_feature_bc_matrix")) : Directory provided does not exist Calls: data.frame -> Read10X Execution halted Work dir: 

Now I'm running SCALPEL with Fastq read file, It's running well.. So I wanted to ask Is there a standardized format for samplesheet.csv file to provide BAM files as input?

[mschilli87]

@Franzx7: The 10X example describes how to specify CellRanger output directorys:

Each row in the samplesheet.csv should contain the following columns, in order, using absolute paths (starting from the root or project directory) for all files and directories:

1. Sample Name: Sample name (…as used by CellRanger).
2. R1 FASTQ Path: The absolute path to the R1 FASTQ file for the sample.
3. R2 FASTQ Path: The absolute path to the R2 FASTQ file for the sample.
4. CellRanger Output Directory Path: The absolute path to the CellRanger output directory for the sample (containing the indexed BAM file).

The Drop-seq example describes how to specify a BAM file along with a DGE count matrix:

Each row in the samplesheet.csv should contain the following columns, in order, using absolute paths (starting from the root or project directory) for all files and directories:

1. Sample Name: Sample name (…as used by CellRanger).
2. R1 FASTQ Path: The absolute path to the R1 FASTQ file for the sample.
3. R2 FASTQ Path: The absolute path to the R2 FASTQ file for the sample.
4. BAM Path: The absolute path to the BAM file for the sample.
5. DGE Count Matrix Path: The absolute path to the DGE count matrix file for the sample.

(note the reference to CellRanger were it should probably mention Drop-seq Tools instead).

The README just mentions 'FASTQ/BAM/CellRanger output' without any details:

Parameter	Description
--samplesheet	CSV with sample names and paths to FASTQ/BAM/CellRanger output

Given that several other issues at some point brought up concerns regarding the sample sheet (e.g. #21 and #24), maybe we should add a detailed description of the format somewhere (ideally in the README or linked there)?