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barCoder: a tool to generate unique, orthogonal genetic tags for qPCR detection
Casey B Bernhards1,2, Matthew W Lux1, Sarah E Katoski1
1U.S. Army Combat Capabilities Development Command Chemical Biological Center, Aberdeen Proving Ground, MD, 21010, USA.
BMC Bioinformatics
|March 2, 2021
Summary
Software-guided design of microbial barcodes simplifies strain identification for environmental tracking. The barCoder tool generates unique, functional DNA tags for microbial simulants, improving detection accuracy and efficiency.
Area of Science:
- Bioinformatics
- Molecular Biology
- Microbial Ecology
Background:
- Environmental microbial tracking requires specific detection methods to differentiate target strains from interferents.
- Genomic insertion of DNA barcodes enables strain detection via real-time PCR.
- Manual barcode design is impractical for large-scale applications, necessitating software solutions.
Purpose of the Study:
- To introduce barCoder, a bioinformatics tool for designing sets of uniquely identifiable, chromosomally barcoded microbial strains.
- To automate and optimize the generation of DNA barcode modules for microbial detection.
Main Methods:
- barCoder utilizes target strain genomic sequences and user-defined PCR parameters.
- It generates DNA barcode modules with primer/probe binding sites and spacers.
- Modules are optimized for PCR amplification (melting temperature, G+C content) and unique identification (BLAST analysis).
Main Results:
- barCoder successfully generated 12 barcode modules for Bacillus thuringiensis and 3 for other organisms.
- Generated barcode modules demonstrated specific real-time PCR detection with minimal cross-reactivity.
- The tool facilitates the creation of synthetically barcoded biological simulants.
Conclusions:
- The barCoder algorithm streamlines the creation of barcoded microbial simulants.
- It reduces manual effort in module design and PCR assay optimization.
- It minimizes the generation of non-unique barcode modules, enhancing detection specificity.
