PrimerBac: A searchable database of taxon-specific primers for rapid microbial identification

Species-resolved PCR detection of gut microorganisms is challenged by within-species genomic diversity and sequence conservation among related taxa. We developed PrimerBac (https://primerbac.cn), a searchable database that integrates gut-focused reference collection, RefSeq pan-genome analysis, core-orthogroup marker selection, background screening, and assembly-level ePCR evaluation. The reference intestinal resource contains 1,762 species, and complete pan-genome analyses were obtained for 1,179 species. PrimerBac provides 1,148,644 primer pairs for 991 prokaryotic species and 611,077 pairs for 424 genera. Broad screening of 22,207 RefSeq reference/representative assemblies yielded 68.9% sensitivity. In a non-overlapping temporal panel of 4,115 post-construction assemblies, 57 of 65 evaluable targets were detected (87.7% sensitivity; 95% CI, 77.5-93.6%), with 84.0% hit-defined macro precision. PrimerBac therefore provides a searchable and systematically evaluated resource for targeted microbial identification.