smithlab/spritz repository overview

⁠Spritz

Software for RNA-Seq analysis on Windows, including creating sample-specific proteoform databases from genomic data

Spritz can be downloaded here⁠.

Spritz uses snakemake and Docker to install and run commandline tools for Next-Generation Sequencing (NGS) analysis. These tools include an adapted version of SnpEff⁠ to annotate sequence variations and create an annotated protein database in XML format. The combinatorics of producing full-length proteoforms from these annotations is written in mzLib's VariantApplication class⁠.

⁠Running Spritz with GUI

1. Install Docker Desktop for Windows⁠.

2. Allocate resources to Docker. There are two ways to do this, described in the Spritz wiki:

   1. The recommended method⁠ requires Windows 10 version 2004 and is more robust. Here, we allocate computer resources to Docker like any other program.
   2. The alternate method⁠ is available on all Windows versions but is less robust. Here, we allocate computer resources to Docker using a virtual machine that's packaged with Docker.

3. Launch Spritz.

   Step 1: Input SRA accessions and/or add FASTQ files.

   • SRAs are added with the button indicating single-end or paired-end.
   • FASTQ files must end with *_1.fastq if single-end, and paired-end sequences must have the same filename other than ending with *_1.fastq and *_2.fastq.

   Step 2: Create and customize your Spritz workflow.

   Step 3: Run Spritz!

   

⁠GUI System Requirements

• Environment:
  • Windows 10 recommended
  • .NET Core 3.1⁠
• 24 GB RAM recommended
• The installer (Spritz.msi⁠) only works on Windows.

⁠Running Spritz with commandline

Spritz will also work on the commandline⁠ within a Unix system (Linux, Mac, WSL on Windows).

⁠Test it out! Try constructing the database for U2OS from the paper.

1. Add SRR629563 to the SRA list.

2. Create the Spritz workflow. Select "release-82" and "homo_sapiens."

3. Run Spritz!

Monitor progress in the Information textbox. The final database named final/combined.spritz.snpeff.protein.withmods.xml.gz can be used to search MS/MS with MetaMorpheus⁠ to find variant peptides and proteoforms, possibly with modifications. We recommend performing 1) Calibration, 2) Global PTM Discovery (G-PTM-D), and 3) Search tasks to get the best results.

The final database named final/combined.spritz.snpeff.protein.fasta is generated to contain variant protein sequences, and it may be used in other search software, such as Proteome Discoverer, ProSight, and MASH Explorer.

The final database named final/combined.spritz.snpeff.protein.withdecoys.fasta is ready for use in MSFragger. It is generated to contain variant protein sequences with decoy protein sequences appended.

⁠Citations

If you use this Spritz, please cite:

• Spritz: Cesnik, A. J.; Miller, R. M.; Ibrahim, K.; Lu, L.; Millikin, R. J.; Shortreed, M. R.; Frey, B. L.; Smith, L. M. “Spritz: A Proteogenomic Database Engine.” J. Proteome Res. 2021, 20, 4, 1826–1834. https://pubs.acs.org/doi/abs/10.1021/acs.jproteome.0c00407⁠

This pipeline uses the following tools:

• sra-toolkit: Leinonen, R.; et al. International Nucleotide Sequence Database Collaboration. The Sequence Read Archive. Nucleic Acids Res. 2011, 39 (Database issue), D19-21. https://doi.org/10.1093/nar/gkq1019⁠.
• fastp: Chen, S.; et al. fastp: an ultra-fast all-in-one FASTQ preprocessor. Bioinformatics 2018, 34 (17), i884-i890. https://academic.oup.com/bioinformatics/article/34/17/i884/5093234⁠
• hisat2: Kim, D.; et al. Graph-Based Genome Alignment and Genotyping with HISAT2 and HISAT-Genotype. Nat. Biotechnol. 2019, 37 (8), 907-915. https://doi.org/10.1038/s41587-019-0201-4⁠."
• samtools: Li, H.; et al. The Sequence Alignment/Map format and SAMtools. Bioinformatics 2009, 25 (16), 2078-2079. https://academic.oup.com/bioinformatics/article/25/16/2078/204688⁠.
• GATK: McKenna, A.; et al. The Genome Analysis Toolkit: A MapReduce Framework for Analyzing next-Generation DNA Sequencing Data. Genome Res. 2010, 20 (9), 1297-1303. https://doi.org/10.1101/gr.107524.110⁠.
• SnpEff: Cingolani, P.; et al. A Program for Annotating and Predicting the Effects of Single Nucleotide Polymorphisms, SnpEff: SNPs in the Genome of Drosophila Melanogaster Strain W1118; Iso-2; Iso-3. Fly (Austin) 2012, 6 (2), 80-92. https://doi.org/10.4161/fly.19695⁠.