Modular universal rbcL primers for (meta)barcoding: a systematic case study clarifying the identity of the marine macroalga Corallina armata (Rhodophyta, Corallinales)

The geniculate coralline algal genus Corallina L. (Rhodophyta, Corallinales) presents longstanding challenges in taxonomy and nomenclature. Recent studies have uncovered previously unrecognised diversity within the genus, highlighting the importance of revisiting historical names and type specimens. In this study, we resolve the taxonomic identity of Corallina armata through DNA sequencing of the rbc L and COI gene regions and designate an epitype to stabilise its nomenclature. Phylogenetic analyses consistently grouped Corallina sequences into species-specific clades across both markers. Using population genetic approaches, single nucleotide polymorphism data revealed clear genetic separation of C. armata from related species, supported by hierarchical clustering, based on genetic distance and principal component analysis of allele frequencies. A pairwise permutational analysis of variance confirmed significant genetic differentiation amongst species (p < 0.05). To support broader taxonomic efforts and utilised in this study, we developed modular universal rbc L primer suites for macroalgae, which performed well in silico across the GenBank database, indicating strong potential for international application (inclusive of some microalgae). Taxonomic identification results from the short-read sequences for rbc L (Ochrophyta and Rhodophyta) and COI showed both primer sets enabled species-level identification, although the rbc L primer sets captured less genetic variation than COI. Nonetheless, each gene revealed distinct interspecific relationships, underscoring the value of multi-locus approaches for robust species delimitation in future barcoding and metabarcoding studies.