# DNA Barcoding as a Plant Identification Method
## 10 Discussion  
DNA barcoding, developed approximately twenty years ago, is an approach that has significantly contributed to the development of molecular systematics. DNA barcodes are standardized sequences, ideally unique, coding or non-coding, either from the genome of the organism or from its organelles, that are used to identify/classify an organismal group; in short, the method includes amplification of the DNA barcode, sequencing and comparison with a reference database containing the relevant sequences from different species. This approach has been extremely successful in animals as _COI_ has emerged as a universal DNA barcode that is used for taxa identification. In plants, however, the use a universal DNA barcode has not been achieved so far. Several studies with the aim of characterizing plant DNA barcodes from chloroplast and nuclear genomes have been performed--_ITS_, _rbCL_, _trnH-pspA_, _rpoB_, _rpoCl_, _trnL-trnF_, _psbK-psbl_, and _atpF-atp_. Based on data from single loci, the CBOL Plant Working Group has suggested the use of combinations of DNA barcodes; however, this approach also has not provided a universal combination for all plants; however, for specific organismal groups, specific combinations have been successfully used. The future of DNA barcodes in plants is next-generation sequencing, through the generation of very long sequences--super barcodes, such as the chloroplast genome, are being used successfully, especially to distinguish between close species. Different strategies could be adopted using next-generation techniques to generate super barcodes in plants; the choice of the best solution may consider different aspects. Annotated chromosome-scale whole-genome sequences (WGS) are the ideal data set in super barcodes [71]; however, the quality of plant material does not always allow for this application. Many sources of samples are stored as historical collections like herbaria [72], and the DNA is degraded with fragments shorter than 100 bp [73]; in this context, it is difficult to apply WGS, and it is necessary to move towards the application of short-read sequencing technologies. Researchers have developed and applied different techniques that realize the generation of thousands of markers both in nuclear DNA, as well as from plastid and mitochondrial genes. While GBS (genotyping by sequencing) and RAD (restriction site-associated genotyping) are mostly useful for plant collection management and diversityand evolutionary studies [74], genome skimming and target capture are more suitable for super barcoding [75]. Target capture is a target enrichment sequencing approach that uses specific probes designed to select target loci obtained from libraries of fragmented genomic DNA. The critical aspect of the technique is the achievement of prior knowledge about the target taxon's DNA to define the probes. The advantages are the moderate cost [76] and the possibility to apply it with low-quality DNA [77]. Genome skimming (also known as shallow shotgun sequencing) is a low-depth sequencing technique [71]. With this approach, it is possible--with a low-medium coverage ranging from 5 to 20 X (it is necessary to consider the genome size and ploidy level)--to recover the fraction of the plant genome that is present in multiple copies, such as the mitochondrial and plastid genomes, nuclear ribosomal DNA (nrDNA) sequences, with higher coverage [78]. Currently, several sequencing services are available in Europe, the USA, and Asian countries, and the costs related to NGS applications are about a few hundred euros per sample, making these approaches feasible, even for small laboratories, assuming there is good bioinformatics support for data analysis. In summary, the future of plant DNA barcoding lies in leveraging next-generation sequencing methodologies, particularly with the emergence of \"super barcodes\". Approaches like genome skimming and target capture demonstrate adaptability in addressing challenges posed by degraded DNA in historical collections.