- Phylogenetic analysis of the whole plastomes based on maximum likelihood and Bayesian inference analyses yielded five groups of Artemisia plastomes clustered in the monophyletic subgenus Dracunculus and paraphyletic subgenus Artemisia, suggesting that the whole plastomes can be used as molecular markers to infer the chloroplast haplotypes of Artemisia taxa. - Conclusions: The complete sequences of the Artemisia plastomes are sufficiently polymorphic to be used as super- barcodes for this genus. - It will facilitate the development of new molecular markers and study of the phylogenomic relationships of Artemisia species in the family Asteraceae.. - Full list of author information is available at the end of the article. - is the largest group in the tribe Anthemideae of the family Asteraceae, consisting of ap- proximately 500 species [1, 2]. - Artemisia species are widely distributed in the temperate regions of the Northern Hemisphere, including Europe, Asia, and North America, and a few species are reported from the South- ern Hemisphere [3–5]. - The center of origin and diversification of the genus Ar- temisia is Asia [8]. - Chloroplasts of the plant kingdom arose from a single ancestral cyanobacterium [26]. - The small and relatively constant size, conserved genome structure, and uniparental inheritance of the plastome make it an ideal genetic resource for phylogenetic analysis and mo- lecular identification of higher plants (reviewed in [27]).. - Several variable regions of the plastome have been devel- oped as DNA barcode marker systems to identify taxa.. - This enables application of the plastome as a super-barcode for high-resolution phylogenetic analysis and species identification [31]. - Table 1 Samples and assembly statistics of the Artemisia plastomes. - We aimed to identify variable regions in the plastomes of the Artemisia taxa in East Asia to establish a molecu- lar basis for the development of novel DNA barcode markers that can be widely applicable across the genus Artemisia as well as the family Asteraceae. - Structure and features of the Artemisia plastomes. - These taxa belong to the sections Abrotanum, Absinthium, and Artemisia of the subgenus Artemisia and the sections Dracunculus and Latilobus of the subgenus Dracunculus [5, 6, 11]. - De novo assembly of the Illumina reads using rbcL and rpoC2 of A. - The structural orientation of the LSC, SSC, and IR regions of each assembly was analyzed by compari- son with previously reported Artemisia plastomes. - Comparing with the plastome of Nicotiana tabacum (GenBank accession NC001879), all the Artemisia plastomes had two inver- sions (approximately 22 kb and 3.3 kb in length) in the LSC region that have been reported to be shared by all clades of the Asteraceae family (Fig. - Table 1 Samples and assembly statistics of the Artemisia plastomes (Continued). - Of the total plastomes, protein-coding genes comprised 52.3%. - Identification of polymorphisms in the Artemisia plastomes. - The alignment exhibited high pairwise sequence identities between plastomes of the same section, ranging from 99.2% (section Absinthium) to 99.8% (section Dra- cunculus) in whole plastomes and from 99.7% (section Absinthium) to 99.9% (section Dracunculus) in the protein-coding genes. - 1 A circular gene map of the Artemisia plastomes. - 0.006) in the coding sequences of accD and ycf1 and in the intron of rps16.. - The PI sites of these genes comprised of 367 sites) of the total PI sites in all protein-coding genes. - Of particular interest, accD, encoding the beta-carboxyl transferase subunit of acetyl-CoA carboxylase, and ycf1, encoding Tic214 of the TIC complex, showed lower sequence identity, higher nucleotide diversity, and a larger number of PI sites than the other genes, indicating a high level of sequence divergence. - protein-coding genes, accD and ycf1, were identified as nucleotide diversity hotspots of the Artemisia chloroplast protein-coding genes, and have potential as candidate regions for the development of universal barcode markers.. - sieversiana (GenBank accession MG951499), one SNP in ndhI induces an in-frame pre- mature stop codon, resulting in loss of eight codons at the 3′-end of the open reading frame.. - The likelihood ratio test of the site-specific model in CodeML program validated the evolutionary selection patterns of accD and ycf1. - In accD, six out of the eight positively selected amino acid. - Table 2 DNA polymorphisms identified in the 32 Artemisia plastomes Structural. - 2 Sliding window test of nucleotide diversity ( π ) in the multiple alignments of the 32 Artemisia plastomes. - Table 3 Evolutionary characteristics of 17 highly diverged protein-coding genes in the Artemisia plastome. - Repetitive sequences in the Artemisia plastomes. - We investigated simple sequence repeats (SSRs) and long sequence repeats (LSRs) in the multiple alignment of the 32 Artemisia whole plastomes (Table 4 and Fig. - Approximately of 431 loci) of SSR loci were in the protein-coding sequences. - In the Artemisia plastomes, di- and trinucleotide repeats were the most frequent, accounting for 72.2 and 18.4% of the total SSRs, respectively (Fig. - less than 15 bp in length, which accounted for 81% of the total LSRs (Additional file 2: Table S7).. - Phylogenetic analysis and delimitation of the Artemisia plastomes. - Using 80 protein-coding genes and the complete plas- tome sequences, we performed phylogenetic analyses of the 32 Artemisia plastomes. - The topology of the max- imum likelihood (ML) and Bayesian inference (BI) trees based on protein-coding genes was nearly identical (Fig. - The plastomes of the subgenus Dracunculus clustered to- gether in monophyletic clade VI whereas those of the sub- genus Artemisia were clustered into four paraphyletic clades (clades I − III and V). - 3 Box-and-whisker plots of the Ka/Ks ( ω ) values of highly diverged protein-coding genes in the Artemisia plastomes. - The pairwise ω values of 14 protein-coding genes in the 32 Artemisia plastomes were calculated and plotted. - Molecular markers for Artemisia and the Asteraceae species A comparative sequence analysis revealed that accD and ycf1 are highly polymorphic in the Artemisia plastomes;. - For accD, the 928 bp un-gapped alignment of the 1 kb region flanking the polymorphic hotspot was investigated as a molecular marker. - For ycf1, we evaluated the potential utility of the Asteraceae ycf1 locus by reconstructing a phylogenetic tree based on the coding sequences of 211 ycf1 genes from Asteraceae whole plastomes in the NCBI nucleotide database. - in agreement with the tribe level taxonomic classification of the Asteraceae (Add- itional file 1: Fig. - 4 Frequency of repetitive sequences in the Artemisia plastomes. - Table 4 Simple sequence repeats and long sequence repeats identified in the 32 Artemisia plastomes. - 5 Phylogenetic tree of Artemisia taxa based on 80 non-redundant protein-coding genes of the plastome. - Colored lines and braces at the right of the tree indicate section and subgenus names of Artemisia. - Table 5 Nucleotide diversity and discriminatory power of the Artemisia chloroplast markers. - The combin- ation of accD and ycf1b (accD-1 k + ycf1b) increased the discriminatory power of the marker. - The topology of the ML tree based on accD-1 k + ycf1b was similar to that of whole plastomes (Additional file 1: Fig. - The Asteraceae ycf1 sequences at the 3′-end of the ycf1bF pri- mer binding site [30] had five to six nucleotide mis- matches. - In contrast, the primer binding sites of ycf1bR as well as of the forward and reverse primers of accD-1 k were mostly conserved. - Considering the consensus se- quences, we designed and optimized primer pairs of the accD-1 k + ycf1b marker for Asteraceae species (Table 6).. - the ycf1b-Asteraceae-F/R primer pair in the Asteraceae family (Additional file 2: Table S8) compared to 0% for the ycf1bF/R primer pair [30]. - However, insufficient sequence polymorphism of the markers and the frequent polyploidy of Artemisia hin- dered their use for species identification. - In the previous comparative plastome analysis of the 11 Artemisia species, intergenic spacer regions, including ccsA-ndhD, trnH- psbA, ndhG-ndhI, rps18-rpl20, and rps15-ycf1, were iden- tified as mutational hotspots. - However, these loci have not been analyzed in the wide range of taxa of the Asteraceae family [40]. - of the whole plastome as a super-barcode is import- ant. - The Artemisia plastomes showed structural character- istics and genetic properties typical of the angiosperm plastome. - selengensis [38] whereas the 5′-region of the ycf1-cod- ing sequence was suggested to be a plastid barcode for land plants [30]. - Moreover, we described the utilization of accD and ycf1 as molecular markers for phylogenetic analysis of the Asteraceae family. - As shown in the ML tree based on 211 Asteraceae ycf1 sequences, ycf1 enabled tribe level resolution of the family Asteraceae (Additional file 1: Fig. - and analyses of conserved orthologs in the Asteraceae. - There was no difference in the tree topology between the ML and BI analyses and there was robust support for most clades, suggesting the validity of the relationships among clades and taxa (Fig. - Interestingly, both phylogenetic trees strongly supported (100%) the monophyletic group of the subgenus Dracun- culus consisting of A. - Molecular markers developed from nuclear genome should provide additional information to distinguish those taxa harbor- ing the same plastome and to create reliable phylogenies of the genus Artemisia.. - A total of 19 individual samples representing 18 taxa of the genus Artemisia (A. - Details of the plant samples are presented in Table 1. - With the draft assemblies, orientation of the SSC was determined according to its structural order in the plastome of A. - Circular gene maps of the Artemisia plastomes were generated using OGDRAW v.1.2 [67]. - Ka and Ks values of the coding sequences were determined using the yn00 program in the PAML v4.9i package [71]. - To choose appropriate outgroup taxa, the species from sev- eral tribes of the Asteraceae, including Cynara humilis in Cardueae, Helianthus annuus in Helentheae, Aster spathulifolius in Astereae, and Chrysanthemun boreale and C. - Comparison of the IR border regions of the Asteraceae plastomes. - Multiple alignments of accD coding sequences in the 32 Artemisia plastomes showing hotspots of nucleotide sequence diversity. - Colored lines and braces at the right of the tree indicate section and subgenus names of Artemisia, respectively, that include taxa. - A ML tree of ycf1 in the Asteraceae family. - A ML tree of accD in the Asteraceae family. - Sample information and statistics of the Illumina PE sequence data of Artemisia taxa. - Gene contents of the Artemisia plastomes. - Pairwise nucleotide similarity matrix of the 32 Artemisia plastomes. - Polymorphic LSRs identified in the 32 Artemisia plastomes. - In silico PCR analysis of the accD- Asteraceae and ycf1b-Asteraceae markers.. - Authors appreciate parataxonomists of The Society for Korean Peninsula Plants (SKPP) who helped collecting Artemisia samples from Korea. - The funding bodies played no role in the design of the study and collection, analysis, and interpretation of data and in writing the manuscript.. - Generic monograph of the Asteraceae- anthemideae. - The Biology and Chemistry of the Compositae. - A systematic study of the genus Artemisia (Asteraceae) in Korea [doctoral thesis]. - The genus Artemisia and ITS allies: phylogeny of the subtribe Artemisiinae (Asteraceae, anthemideae) based on nucleotide sequences of nuclear ribosomal DNA internal transcribed spacers (ITS). - Origin and evolution of the chloroplast division machinery. - The evolution of the plastid chromosome in land plants: gene content, gene order, gene function. - Complete chloroplast genome sequence and phylogenetic analysis of the medicinal plant Artemisia annua. - Two chloroplast DNA inversions originated simultaneously during the early evolution of the sunflower family (Asteraceae). - Tribal interrelationships of the Asteraceae. - Utility of the trnH-psbA intergenic spacer region and its combinations as plant DNA barcodes: a meta-analysis. - Potential functional replacement of the plastidic acetyl-CoA carboxylase subunit (accD) gene by recent transfers to the nucleus in some angiosperm lineages
Xem thử không khả dụng, vui lòng xem tại trang nguồn hoặc xem
Tóm tắt