PUPpy

PUPpy designs microbe- and group-specific primers within a given microbial community to enable detection and absolute quantification of microbial DNA at substrain-level resolution.

Key Features:

• Automated primer design: PUPpy fully automates the design of taxon-specific primers using only coding sequence files of community members as input.
• Substrain-level specificity: PUPpy-designed primers achieve specificity below the strain level for detecting individual microbes within defined communities.
• Group-specific detection: PUPpy-designed primers enable detection of groups of bacteria in undefined communities, such as a gut bacterial family in complex stool microbiota.
• Absolute quantification support: PUPpy-designed primers enable absolute quantification of microbial abundance via droplet digital PCR.
• Benchmarked specificity: Taxon-specific primers designed with PUPpy showed 100% specificity to intended targets without unintended amplification in tested communities spanning genetic similarities from different phyla to substrains.

Scientific Applications:

• Defined community profiling: PUPpy enables selective detection and absolute quantification of individual microbes in defined bacterial communities with higher resolution and accuracy than short-read 16S rRNA and shotgun sequencing.
• Complex microbiota analysis: PUPpy-designed primers can detect specific bacterial groups, such as a gut bacterial family, in complex stool microbiota samples.
• Ecological interaction studies: PUPpy supports characterization of bacterial communities in both in vitro and complex microbiota settings to uncover hidden ecological interactions within microbial ecosystems.

Methodology:

PUPpy is implemented in Python and designs phylogenetically unique primers from coding sequence files of community members.