Conserved single-nucleotide polymorphisms (SNPs) which characterize the allelic profile of the major epidemiological lineage ST-21 were identified from the alleles within the current Campylobacter jejuni multilocus sequence typing (MLST) database. Allelic discrimination assays were designed for the detection of SNPs, enabling rapid strain profiling for clonal complex ST-21. This method is suitable for epidemiological investigations and is complementary to full MLST.