Strategies for molecular authentication of herbal products: from experimental design to data analysis

Molecular herbal authentication has gained worldwide popularity in the past decade. DNA-based methods, including DNA barcoding and species-specific amplification, have been adopted for herbal identification by various pharmacopoeias. Development of next-generating sequencing (NGS) drastically increased the throughput of sequencing process and has sped up sequence collection and assembly of organelle genomes, making more and more reference sequences/genomes available. NGS allows simultaneous sequencing of multiple reads, opening up the opportunity of identifying multiple species from one sample in one go. Two major experimental approaches have been applied in recent publications of identification of herbal products by NGS, the PCR-dependent DNA metabarcoding and PCR-free genome skimming/shotgun metagenomics. This review provides a brief introduction of the use of DNA metabarcoding and genome skimming/shotgun metagenomics in authentication of herbal products and discusses some important considerations in experimental design for botanical identification by NGS, with a specific focus on quality control, reference sequence database and different taxon assignment programs. The potential of quantification or abundance estimation by NGS is discussed and new scientific findings that could potentially interfere with accurate taxon assignment and/or quantification is presented.