Introduction

buildrnaseqbundle is a bioinformatics pipeline that can be used to create bundles of input data (genome files, annotations, indexes etc) for RNA sequencing analyses with nf-core/rnaseq, using the guidelines for GTEx v10.

1. Get the input fasta and gtf files
   1. Prepre the reference genome file
      1. Get the GRCh38 reference genome FASTA from Broad Institute (GRCh38)
      2. Remove the ALT, HLA, and Decoy contigs from the reference genome FASTA
   2. Prepre the gene annotation file (gunzip)
      1. Get the Gencode v.XX annotation of choice (Gencode)
   3. Prepare the ERCC92 spike-in data (7za, sed)
      1. Get the Thermofisher ERCC spike-in data (ERCC92 spike-in)
      2. Patch the ERCC92 fasta file for compatibility with RNA-SeQC/GATK
2. Combine the Gencode and ERCC GTF annotation files (cat)
3. Create the STAR index (STAR, tar)
4. Create the RSEM index (RSEM, tar)
5. Create the combined fasta fai and dict files for GATK/Picard (samtools, gatk4)

Usage

Note If you are new to Nextflow and nf-core, please refer to this page on how to set-up Nextflow. Make sure to test your setup with -profile test before running the workflow on actual data.

You can run the pipeline using:

nextflow run nf-core/buildrnaseqbundle \
   -profile <docker/singularity/.../institute> \
   --spliceJunctionOverhang    100 \
   --outdir                    <OUTDIR>

Warning: Please provide pipeline parameters via the CLI or Nextflow -params-file option. Custom config files including those provided by the -c Nextflow option can be used to provide any configuration except for parameters; see docs.

For more details and further functionality, please refer to the usage documentation and the parameter documentation.

Pipeline output

The pipeline creates an output folder with the same output as the GTEx v10 analysis, as well as the input for the rnaseq pipeline input.

nf-test-rnaseq-hg38-gencode.v44-bundle/
├── gencode.v44.annotation_genes_collapsed_only_patched_ERCC92.gtf
├── Homo_sapiens_assembly38_noALT_noHLA_noDecoy_patched_ERCC92.dict
├── Homo_sapiens_assembly38_noALT_noHLA_noDecoy_patched_ERCC92.fasta
├── Homo_sapiens_assembly38_noALT_noHLA_noDecoy_patched_ERCC92.fasta.fai
├── pipeline_info/
├── rsem_reference_GRCh38_gencode44_ercc.tar.gz
├── star_rsem_index/
└── STARv2710a_genome_GRCh38_noALT_noHLA_noDecoy_ERCC_v44_oh100.tar.gz

Current versions

	Version
Genome	GRCh38
GENCODE	v44
python	3.10.2
samtools	1.17
gatk4	4.4.0.0
STAR	2.7.10a
RSEM	1.3.3

Credits

nf-core/buildrnaseqbundle was originally written by Evangelos (Vangelis) Theodorakis.

Contributions and Support

If you would like to contribute to this pipeline, please see the contributing guidelines.

For further information or help, don't hesitate to get in touch on the Slack #buildrnaseqbundle channel (you can join with this invite).

Citations