Evaluation of DNA barcode candidates for the discrimination of Artemisia L.

Abstract

Because of the very similar morphologies and wide diversity of Artemisia L. varieties, they are difficult to identify, and there have been many arguments about the systematic classification Artemisia L., especially concerning the division of species. DNA barcode technology is used to rapidly identify species based on standard short DNA sequences. To evaluate seven candidate DNA barcodes (ITS, ITS2, psbA-trnH, rbcL, matK, rpoB, and rpoC1) regarding their ability to identify closely related species of the Artemisia genus in Xinjiang. The corresponding PCR amplification efficiency, detectable genetic divergence, identification efficiency and phylogenetic tree were assessed. We found that the internal transcribed spacer (ITS) region exhibited the highest interspecific divergence, which was significantly higher than the observed intraspecific variation and showed the highest identification efficiency, followed by ITS2, psbA-trnH and, finally, rpoB. matK, rbcL, and rpoC1 performed poorly in this evaluation. ITS, ITS2, and psbA-trnH were able to perfectly identify the tested species Artemisia annua, A. absinthium, A. rupestris, A. tonurnefortiana, A. austriaca, A. dracunculus, A. vulgaris, and A. macrocephala. Therefore, we propose the ITS, ITS2, and psbA-trnH regions as promising DNA barcodes for the closely related species of Artemisia L. in Xinjiang.

Comparative chloroplast genome analysis of Artemisia (Asteraceae) in East Asia: insights into evolutionary divergence and phylogenomic implications

“…We anticipate that successful PCR amplification of these intergenic spacers and other polymorphic LSR loci will enable development of a single DNA marker for genotype identification below the species level. Meanwhile, consistent with a report of selected chloroplast genes in a limited number of Artemisia species in China [21], most protein-coding genes had low sequence divergence, with the exception of accD, ycf1, and rps16. These genes contained nucleotide diversity hotspots in exon (accD and ycf1) or intron (rps16) regions that were under weak positive selection or relaxed selective constraints.…”

“…However, these loci have not been analyzed in the wide range of taxa of the Asteraceae family [40]. Additionally, known core chloroplast barcodes, such as rbcL, matK, rpoB and rpoC1, had low discriminatory power for classification of Artemisia taxa [21][22][23]. Therefore, discovery of novel barcodes with high discriminatory power or use Universal barcode primer set designed for angiosperm [30] of the whole plastome as a super-barcode is important.…”

“…Recently, the subgenus Pacifica, including Hawaiian species, was recognized by nuclear ribosomal (ITS and ETS) and chloroplast (trnL-F and psbA-trnH) markers [20]. However, the resolution of these markers was insufficient to resolve taxonomic issues at the species level due to high sequence similarity of closely related taxa presumably caused by rapid radiation and hybridization [21][22][23][24]. Therefore, development of novel DNA markers or barcodes for investigation of Artemisia is an important challenge.…”

Optimisation of artemisinin and scopoletin extraction from Artemisia annua with a new modern pressurised cyclic solid–liquid (PCSL) extraction technique

“…Approximately 200 g of silica gel‐dried leaves of two samples of A. annua were ground in liquid nitrogen. Total genomic DNA was extracted in CTAB buffer . Polymerase chain reaction (PCR) amplification of the ribosomal internal transcribed spacers, ITS1 and ITS2, was performed as described in Aceto et al .…”

“…Total genomic DNA was extracted in CTAB buffer. 23,24 Polymerase chain reaction (PCR) amplification of the ribosomal internal transcribed spacers, ITS1 and ITS2, was performed as described in Aceto et al 25 The amplification products were directly sequenced and run on the 310 ABI Genetic Analyser (Applied Biosystems).…”