Genetic authentication by RFLP versus ARMS? The case of Moldavian Dragonhead (Dracocephalum moldavica L.)
Horn T, Völker J, Rühle M, Häser A, Jürges G, Nick P
2014, Eur Food Sci Technol 238, 93-104
Moldavian Dragonhead (Dracocephalum moldavica L.), due to its pleasant Lemon Scent and medical effects has acquired increasing impact as functional food. The high diversity within the genus, limited supply not keeping pace with the growing demand, the morphological similarity with other Labiatae, and trading under the common name Turkish Melissa invite adulteration by surrogate species. We have developed several verified reference accessions of D. moldavica L. along with potential surrogate species to compare different approaches of authentication, also in commercial samples. We report on three strategies of authentication – a microscopical method, based on the relative size of epidermal pavement cells versus palisade cells, and two strategies of genetic authentication based on barcoding marker large subunit of ribulose-1,5-bisphosphate carboxylase oxygenase (rbcL). We can detect single-nucleotide exchanges between D. moldavica L. and the potential surrogate species Melissa officinalis L. and Nepeta cataria L. by restriction fragment length polymorphism (RFLP), and we show that we use this to verify the presence of D. moldavica even in dried and highly fragmented mixtures from commercial samples. We further develop a third strategy derived from the so called Amplification Refractory Mutation System (ARMS), based on multiplex PCR of the rbcL marker upon addition of specifically designed intermediate primers that will generate a diagnostic second band in case of D. moldavica L., but not for the surrogate species. We demonstrate that this ARMS approach is superior over the RFLP strategy, because it safeguards against experimental fluctuations, can unequivocally verify the presence of D. moldavica in commercial samples declaring its presence, and yields a clear outcome in a one-step protocol.