Applying DNA Barcodes to Identify Closely Related Species of Ferns: A Case Study of the Chinese Adiantum (Pteridaceae)

Abstract

DNA barcoding is a fast-developing technique to identify species by using short and standard DNA sequences. Universal selection of DNA barcodes in ferns remains unresolved. In this study, five plastid regions (rbcL, matK, trnH-psbA, trnL-F and rps4-trnS) and eight nuclear regions (ITS, pgiC, gapC, LEAFY, ITS2, IBR3_2, DET1, and SQD1_1) were screened and evaluated in the fern genus Adiantum from China and neighboring areas. Due to low primer universality (matK) and/or the existence of multiple copies (ITS), the commonly used barcodes matK and ITS were not appropriate for Adiantum. The PCR amplification rate was extremely low in all nuclear genes except for IBR3_2. rbcL had the highest PCR amplification rate (94.33%) and sequencing success rate (90.78%), while trnH-psbA had the highest species identification rate (75%). With the consideration of discriminatory power, cost-efficiency and effort, the two-barcode combination of rbcL+ trnH-psbA seems to be the best choice for barcoding Adiantum, and perhaps basal polypod ferns in general. The nuclear IBR3_2 showed 100% PCR amplification success rate in Adiantum, however, it seemed that only diploid species could acquire clean sequences without cloning. With cloning, IBR3_2 can successfully distinguish cryptic species and hybrid species from their related species. Because hybridization and allopolyploidy are common in ferns, we argue for including a selected group of nuclear loci as barcodes, especially via the next-generation sequencing, as it is much more efficient to obtain single-copy nuclear loci without the cloning procedure.

Using near-infrared spectroscopy to discriminate closely related species: A case study of neotropical ferns

“…The high predictive power of FT-NIR at discriminating fern species, presented here, is superior to that observed for this lineage using a single region of two combined DNA barcodes, for which the best performance was 75 % correct predictions (Li et al 2011; Wang et al 2016). Identifying ferns and other plant groups using DNA barcoding is also an expensive technique and different lineages require specific combinations of molecular markers, which can make this technique complicated (Li et al 2011; Lima et al 2018).…”

Constructing a DNA barcode reference library for southern herbs in China: A resource for authentication of southern Chinese medicine

“…Although ITS2 has many advantages, it does not work as well in some taxa of ferns. Low PCR efficiency has become an obvious shortcoming based on our study and other studies [ 12 , 32 ]. Heterogeneity is also a problematic issue for ITS2 as a consequence of concerted evolution [ 25 ].…”

Evaluation of cpDNA Barcodes in Selected Medicinal Plants of Mt. Arayat National Park, Pampanga, the Philippines

“…Since the best currently available 'universal' primer pair of matK (3F/1R) which produces 70% PCR and sequencing success in angiosperms on diverse sample sets, it was recommended to use a secondary primer pair (390F/1326R) which can increase amplification and sequencing success by another 10%. 15 While in the study of Wang et al (2016) in the DNA barcoding of ferns, Chinese Adiantum L., the amplification of trnL-F was discovered to be relatively difficult because of the universal presence of mononucleotide repeats in the intergenic spacer that affected sequence quality and reduced the universality of this marker. 24.…”

“…15 While in the study of Wang et al (2016) in the DNA barcoding of ferns, Chinese Adiantum L., the amplification of trnL-F was discovered to be relatively difficult because of the universal presence of mononucleotide repeats in the intergenic spacer that affected sequence quality and reduced the universality of this marker. 24.…”