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mapo9 committed May 16, 2024
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50 changes: 47 additions & 3 deletions docs/usage.md
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Expand Up @@ -42,13 +42,13 @@ nextflow run nf-core/airrflow \
A typical command to run the pipeline from **single cell raw fastq files** is:

```bash
nextflow run nf-core/airrflow -r dev \
nextflow run nf-core/airrflow \
-profile <docker/singularity/podman/shifter/charliecloud/conda/institute> \
--mode fastq \
--input input_samplesheet.tsv \
--library_generation_method sc_10x_genomics \
--reference_10x reference/refdata-cellranger-vdj-GRCh38-alts-ensembl-5.0.0.tar.gz \
--outdir ./results
--outdir results
```

A typical command for running the pipeline departing from **single-cell AIRR rearrangement tables or assembled bulk sequencing fasta** data is:
Expand Down Expand Up @@ -123,7 +123,7 @@ If you wish to share such profile (such as upload as supplementary material for

## Input samplesheet

### Fastq input samplesheet (bulk sequencing)
### Fastq input samplesheet (bulk AIRR and bulk/sc RNA sequencing)

The required input file for processing raw BCR or TCR bulk targeted sequencing data is a sample sheet in TSV format (tab separated). The columns `sample_id`, `filename_R1`, `filename_R2`, `subject_id`, `species`, `tissue`, `pcr_target_locus`, `single_cell`, `sex`, `age` and `biomaterial_provider` are required. An example samplesheet is:

Expand Down Expand Up @@ -511,6 +511,50 @@ nextflow run nf-core/airrflow -r dev \
- The 10xGenomics reference can be downloaded from the [download page](https://www.10xgenomics.com/support/software/cell-ranger/downloads)
- To generate a V(D)J segment fasta file as reference from IMGT one can follow the [cellranger docs](https://support.10xgenomics.com/single-cell-vdj/software/pipelines/latest/advanced/references#imgt).


## Supported unselected RNA-seq based methods

nf-core/airrflow supports unselected bulk or single-cell RNA-seq fastq files as input. [TRUST4](https://github.com/liulab-dfci/TRUST4) is used to extract TCR/BCR sequences from these files. The resulting AIRR tables are then fed into airrflow's Immcantation based workflow. <br>
To use unselected RNA-seq based input, specify `--library_generation_method trust4`.

### Bulk RNA-seq

A typical command to run the pipeline from **bulk RNA-seq fastq files** is:

```bash
nextflow run nf-core/airrfow \
-profile <docker/singularity/podman/shifter/charliecloud/conda/institute> \
--mode fastq \
--input input_samplesheet.tsv \
--library_generation_method trust4 \
--coord_fasta reference/IMGT+C.fa \
--outdir results
```

### Single-cell RNA-seq

A typical command to run the pipeline from **single-cell RNA-seq fastq files** is:

```bash
nextflow run nf-core/airrfow \
-profile <docker/singularity/podman/shifter/charliecloud/conda/institute> \
--mode fastq \
--input input_samplesheet.tsv \
--library_generation_method trust4 \
--umi_position R1 \
--read_format bc:0:15,um:16:27
--coord_fasta reference/IMGT+C.fa \
--outdir results
```

* If UMI's are present, the read containing them must be specified using the `--umi_position` parameter.
* The `--read_format` parameter can be used to specify the Barcode and UMI position within the reads (see TRUST4 [docs](https://github.com/liulab-dfci/TRUST4?tab=readme-ov-file#10x-genomics-data-and-barcode-based-single-cell-data))

#### Reference file

TRUST4 requires a reference. This can provided using the `--coord_fasta` parameter.
The reference fasta can be downloaded from IMGT and created using [TRUST4](https://github.com/liulab-dfci/TRUST4?tab=readme-ov-file#build-custom-vjc-gene-database-files-for--f-and---ref)

## Core Nextflow arguments

:::note
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4 changes: 2 additions & 2 deletions nextflow_schema.json
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Expand Up @@ -487,9 +487,9 @@
"fa_icon": "fab fa-pagelines"
},
"rnaseq_based_analysis_options": {
"title": "RNA seq based analysis options",
"title": "Unselected RNA-seq based analysis options",
"type": "object",
"description": "Options specific for raw RNA seq input.",
"description": "Options specific for raw unselected RNA-seq input.",
"default": "",
"properties": {
"barcode_read": {
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