Pannagram

Overview

Pannagram is a toolkit for building reference-free linear pangenome alignments and analyzing genomic polymorphisms.
It consists of a command-line interface (CLI) for alignment construction, feature extraction, and sequence search, and an R library for downstream analysis and visualization.

Key capabilities:

• Reference-free pangenome alignment
• SNP and structural variant detection
• Mobile element family discovery
• Search for sequences in genomes
• Annotation liftover between genomes
• Visualization and sequence analysis

Documentation can be found at Pannagram-page.

Quick Installation

Clone the repository and create the conda environment:

git clone https://github.com/<user>/pannagram.git
cd pannagram
conda env create -f pannagram.yml
conda activate pannagram
./user.sh
./verify_installation.sh  # Verify the successful installation

Quick Start

The typical workflow consists of two steps:

1. Build the pangenome alignment
2. Call genomic features from the alignment

Before running the example, set the following variables (preferably absolute paths) in the command line:

• PATH_GENOMES – directory containing input genome FASTA files
• PATH_PROJECT – directory where the project output will be stored

Reference-Free Pangenome Alignment

Run the following command to perform a reference-free pangenome alignment:

pannagram  -path_genomes ${PATH_GENOMES} \
           -path_project ${PATH_PROJECT} \
           -cores 8

Feature Calling

After the alignment step is complete, run the feature-calling module to identify all available genomic features:

features  -path_project ${PATH_PROJECT} \
          -synteny \
          -consensus \
          -snp \
          -snp_pi \
          -sv \
          -sv_families \
          -cores 8

All results will be saved under ${PATH_PROJECT} after both steps are complete:

PATH_PROJECT/  
├── features/     ← main analysis outputs  
└── plots/        ← visualizations and figures  

A detailed description of all output files and their formats is available in the documentation under Getting Started → Output Data.

Pannagram R Library

In your R session, load the library:

library(pannagram)

Pannagram R library provides functions for:

• working with FASTA files
• annotation liftover
• extracting specific pangenome regions as multiple sequence alignments
• ORF finding and visualization
• dot plots
• multiple sequence alignment visualization
• pangenome plots

For detailed documentation, visit the Pannagram-page.

Citation

If you use Pannagram, please cite:

• Pannagram: unbiased pangenome alignment and Mobilome calling
  Anna A. Igolkina et al., bioRxiv, 2025. Link

To explore Pannagram applications, we recommend:

• A comparison of 27 Arabidopsis thaliana genomes and the path toward an unbiased characterization of genetic polymorphism
  Anna A. Igolkina et al., Nature Genetics, 2025. Link

Acknowledgements

Development:

• Anna Igolkina - Lead Developer and Project Initiator
• Alexander Bezlepsky - Assistant

Testing:

• Anna Igolkina: Lead Tester
• Anna Glushkevich: Testing the alignment on A. lyrata genomes
• Elizaveta Grigoreva: Testing the alignment on A. thaliana and A. lyrata genomes
• Jilong Ma: Testing the SV-graph on spider genomes
• Alexander Bezlepsky: Testing the Pannagram's functionality on Rhizobial genomes
• Gregoire Bohl-Viallefond: Testing the annotation converter on A. thaliana alignment

Resources:

• Parallel Processing Tool: O. Tange (2018): GNU Parallel 2018, ISBN 9781387509881, DOI https://doi.org/10.5281/zenodo.1146014.