Efficient coralline algal psb A mini barcoding and High Resolution Melt (HRM) analysis using a simple custom DNA preparation

Coralline algaeform extensive maerland rhodolithhabitats that support a rich biodiversity. Calcium carbonate harvesting as well as trawlingactivities threatens this ecosystem. Eleven species were recorded so far as maerl-forming in NE Atlantic, but identification based on morphological characters is unreliable. As for most red algae, we now use molecular characters to resolve identification of these taxa. However, obtaining DNA sequences requires time and resource demanding methods. The purpose of our study was to improve methods for achieving simple DNA extraction, amplification, sequencing and sequence analysis to allow robust identification of maerlspecies and other coralline algae. Our novel and easy DNA preparation method for coralline algaewas of sufficient quality for qPCR amplification and sequencing of all 47 tested samples. The new psbA qPCR assay successfully amplified a 350 bp fragment identifyingsix species and uncovering two new Operational Taxonomic Units. Molecular results were corroborated with anatomical examination using i.e. scanning electron microscopy. Finally, the qPCR assay was coupled with High Resolution Meltanalysis that successfully differentiated the closely related species Lithothamnionerinaceum and L. cf. glaciale. This DNA preparation and qPCR technique should vitalize coralline research by reducing time and cost associated with molecular systematics.