Primerdiffer: a python command-line module for large-scale primer design in haplotype genotyping

Motivation: Primer design is a routine practice for modern molecular biology labs. Bioinformatics tools like primer3 and primer-blast have standardized the primer design for a specific region. However, large-scale primer design, especially for genome-wide screening, is still a labor-intensive job for most wet-lab researchers using these pipelines.Results: Here, we present the primerdiffer pipeline, which can be used to batch design primers that differentiate haplotypes on a large scale with precise false priming checking. This command-line interface (CLI) pipeline includes greedy primer search, local and global in silico PCR-based false priming checking, and automated best primer selection. The local CLI application provides flexibility to design primers with the user's own genome sequences and specific parameters. Some species-specific primers designed to genotype the hybrid introgression strains from Caenorhabditis briggsae and Caenorhabditis nigoni have been validated using single-worm PCR. This pipeline provides the first CLI-based large-scale primer design tool to differentiate haplotypes in any targeted region.

Automated summary

The primerdiffer pipeline is introduced as a large-scale primer design tool that can differentiate haplotypes with precise false priming checking. It includes steps like greedy primer search, in silico PCR-based false priming checking, and automated best primer selection. The pipeline provides a command-line interface and flexibility for users to design primers with their own genome sequences and specific parameters.