Chloroplast Non-coding Regions Research Articles

Molecular evolution of chloroplast DNA in fig ( Ficus carica L.): Footprints of sweep selection and recent expansion

Here, we report the nucleotide variation in two non-coding regions of the chloroplast DNA (cpDNA) to construct a possible evolutionary scenario in Ficus carica L. Our results suggest the occurrence of haplotypic and nucleotide diversity with a large variation level of chloroplast non-coding regions. Furthermore, our results demonstrated an explicit rejection of the null hypothesis that within F. carica the intron trnL and the spacer trnL- trnF evolved under a strictly neutral model of molecular evolution. Although, recent population expansion could serve as one alternative explanation for the detected excess of singleton, our results imply a positive selection and the genetic hitchhiking effect is unlikely. Parameters performed supported scenario of sweep selection and recent expansion of F. carica across Tunisia. Our results indicate that both positive selection and demographic histories have jointly contributed to the observed patterns of nucleotide diversity and haplotypes structure. Based on the results, we characterize the fig resources and provide several suggestions for effective conservation and improvement programs.

Large multi-locus plastid phylogeny of the tribe Arundinarieae (Poaceae: Bambusoideae) reveals ten major lineages and low rate of molecular divergence

The temperate bamboos (tribe Arundinarieae) are notorious for being taxonomically extremely difficult. China contains some of the world’s greatest diversity of the tribe Arundinarieae, with most genera and species endemic. Previous investigation into phylogenetic relationships of the temperate bamboos revealed several major clades, but emphasis on the species-level relationships among taxa in North America and Japan. To further elucidate relationships among the temperate bamboos, a very broad sampling of Chinese representatives was examined. We produced 9463 bp of sequences from eight non-coding chloroplast regions for 146 species in 26 genera and 5 outgroups. The loci sequenced were atpI/H, psaA-ORF170, rpl32- trnL, rpoB- trnC, rps16- trnQ, trnD/T, trnS/G, and trnT/L. Phylogenetic analyses using maximum parsimony and Bayesian inference supported the monophyly of Arundinarieae. The two major subtribes, Arundinariinae and Shibataeinae, defined on the basis of different synflorescence types, were indicated to be polyphyletic. Most genera in this tribe were confirmed to be paraphyletic or polyphyletic. The cladograms suggest that Arundinarieae is divided into ten major lineages. In addition to six lineages suggested in a previous molecular study (Bergbamboes, the African alpine bamboos, Chimonocalamus, the Shibataea clade, the Phyllostachys clade, and the Arundinaria clade), four additional lineages were recovered in our results, each represented by a single species: Gaoligongshania megalothyrsa, Indocalamus sinicus, Indocalamus wilsonii, Thamnocalamus spathiflorus. Our analyses also indicate that (1) even more than 9000 bp of fast-evolving plastid sequence data cannot resolve the inter- and infra-relationships among and within the ten lineages of the tribe Arundinarieae; (2) an extensive sampling is indispensable for phylogeny reconstruction in this tribe, especially given that many genera appear to be paraphyletic or polyphyletic. Perhaps the ideal way to further illuminate relationships among the temperate bamboos is to sample multiple nuclear loci or whole chloroplast sequences in order to obtain sufficient variation.

Reassignment of Species of Paraphyletic Junellia s. l. to the New Genus Mulguraea (Verbenaceae) and New Circumscription of Genus Junellia: Molecular and Morphological Congruence

Fil: O'Leary, Nataly Cristina. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Instituto de Botanica Darwinion. Academia Nacional de Ciencias Exactas, Fisicas y Naturales. Instituto de Botanica Darwinion; Argentina

Authentication of the Medicinal Plants in Fabaceae by DNA Barcoding Technique

Fabaceae is the third largest family of flowering plants, with a large number of medicinal plants. However, it is arduous to identify some of the species in this family because of morphological similarity and frequent variation. The DNA barcode, a short DNA sequence originating from the genome, was first investigated for the medicinal plant in Fabaceae. Now we have completed 86 species of medicinal plants in Fabaceae including over 30 genera. Through six candidates, promising markers, four coding (rpoB, rpoC1, rbcL, matK) and two noncoding (ITS, ITS2) chloroplast regions, we identified potential barcodes of the medicinal plants in Fabaceae by comparing DNA barcoding sequences. The results indicated, efficiency of amplification for six candidate DNA barcodes range from 100% (ITS2) to 93% (matk). Intraspecific variation and interspecific variation for six chloroplast regions derived Wilcoxon signed rank tests of divergence showed that rpoC1 was the least discriminatory region, while ITS2 was the potential candidate of medicinal plant barcoding in Fabaceae in our study. Overall, our findings showed that DNA barcoding is an efficient and powerful tool for the identification of the medicinal plants in Fabaceae. Acknowledgements: This work is supported by the International Cooperation Program of Science and Technology (No. 2007DFA30990) and the Special Founding for Healthy Field (No. 200802043).

Inferred Phylogeny in Keckiella (Scrophulariaceae) based on Noncoding Chloroplast and nuclear ribosomal DNA Sequences

Abstract Two non-coding DNA regions from both the chloroplast genome (trnL intron, trnL–trnF IGS, trnT–trnL IGS) and nuclear ribosomal genomes (ITS1 and ITS2) were sequenced in the genus Keckiella and allied genera (Penstemon, Chelone, and Nothochelone) to determine which, if any, of these sequences contained sufficient variability to resolve the seven species of Keckiella. The trnL intron contained almost no signal. However, the trnT–trnL chloroplast IGS region and the nuclear ribosomal sequences were capable, to some extent, of resolving the phylogeny of Keckiella. The nrDNA ITS sequences contained slightly more variability than the chloroplast noncoding region. When combined, these non-coding sequences generated a relatively robust and resolved cladogram. Keckiella rothrockii is clearly indicated as the most basal species of the genus, and the remaining taxa appear to belong to two well-supported clades. The first contains the sister taxa K. antirrhinoides and K. cordifolia. The second contains K. brev...

Chloroplast DNA analysis in Tunisian fig cultivars ( Ficus carica L.): Sequence variations of the trnL-trnF intergenic spacer

We investigated the nucleotide variation of a non-coding, chloroplast DNA (cpDNA) region to infer relationships among Tunisian fig cultivars. In this study, we examine the level of genetic diversity and its distribution using sequences of the trnL and trnF genes intergenic spacer. The non-coding region displays 28 substitution sites. Insertions and deletions involving 6 sites were found. By using the Kimura-2 method, nucleotide sequences have been aligned using the MEGA program to calculate pairwise divergence of trnL-trnF spacer sequences between cultivars. The size of this non-coding region varied from 430 to 464 bases. The relatively high A + T values (63.7–64.4%) of trnL-trnF intergenic spacer in Ficus carica may explain the high proportion of the identified transversions ( t i/ t v = 0.9). These results suggest the occurrence of nucleotide diversity with a large variation level of chloroplast non-coding region. The analysed data illustrate a considerable level of variability in the genetic pool of the local germplasm. In fact, relationships inferred from the cpDNA analysis suggest several clades, which do not show geographical correspondence. Fourteen haplotypes were detected among 20 individuals examined, yielding a haplotype diversity of 0.983 and a high level of nucleotide diversity (0.0100). The observed variation pattern of plastid DNA provides evidence that the fig germplasm has been undergoing rapid expansion. Neutrality tests rejected the neutrality assumption in the total sample. The cytoplasm variability indicates a narrow genetic base in the cultivated common fig. Despite the high level value of the apparent diversity, we may conclude that fig chloroplast genome provides a new conceptual and practical opportunity to evaluate genetic diversity and to identify local cultivars, making it a valuable source to include into potential breeding programs.

Phylogenetics of the Caprifolieae and <I>Lonicera</I> (Dipsacales) Based on Nuclear and Chloroplast DNA Sequences

Abstract Recent phylogenetic analyses of the Dipsacales strongly support a Caprifolieae clade within Caprifoliaceae including Leycesteria, Triosteum, Symphoricarpos, and Lonicera. Relationships within Caprifolieae, however, remain quite uncertain, and the monophyly of Lonicera, the most species-rich of the traditional genera, and its subdivisions, need to be evaluated. In this study we used sequences of the ITS region of nuclear ribosomal DNA and five chloroplast non-coding regions (rpoB–trnC spacer, atpB–rbcL spacer, trnS–trnG spacer, petN–psbM spacer, and psbM–trnD spacer) to address these problems. Our results indicate that Heptacodium is sister to Caprifolieae, Triosteum is sister to the remaining genera within the tribe, and Leycesteria and Symphoricarpos form a clade that is sister to a monophyletic Lonicera. Within Lonicera, the major split is between subgenus Caprifolium and subgenus Lonicera. Within subgenus Lonicera, sections Coeloxylosteum, Isoxylosteum, and Nintooa are nested within the paraph...

Toward a comprehensive understanding of phylogenetic relationships among lineages of Acanthaceae s.l. (Lamiales)

Acanthaceae (Asteridae; Lamiales) include ∼4000 species and encompass a range of morphological diversity, habitats, and biogeographic patterns. Although they are important components of tropical and subtropical habitats worldwide, inadequate knowledge of the family's phylogenetic framework has impeded comparative research. In this study, we sampled all known lineages of Acanthaceae including Andrographideae. Also included were eight of 13 genera whose relationships remain enigmatic. We used sequence data from nrITS and four chloroplast noncoding regions, and parsimony and Bayesian methods of analysis. Results strongly support most aspects of relationships including inclusion of Avicennia in Acanthaceae. Excepting Neuracanthus, newly sampled taxa are placed with strong support; Kudoacanthus is in Justicieae, Tetramerium lineage, and the remaining enigmatic genera are in Whitfieldieae or Barlerieae, and Andrographideae are sister to Barlerieae. This last result is unanticipated, but placement of Andrographideae based on structural characters has been elusive. Neuracanthus is monophyletic but placement relative to (Whitfieldieae (Andrographideae + Barlerieae)) is weakly supported. Many clades have clear morphological synapomorphies, but nonmolecular evidence for some remains elusive. Results suggest an Old World origin with multiple dispersal events to the New World. This study informs future work by clarifying sampling strategy and identifying aspects of relationships that require further study.

Hybrid Origin and Genomic Mosaicism of Dubautia scabra (Hawaiian Silversword Alliance; Asteraceae, Madiinae)

Incongruence among different estimates of species relationships in plants, from different molecules, cytogenetic data, biogeographic data, morphological/anatomical data or other sources, has been used frequently as an indication of introgression, hybrid species origin, or chloroplast (cp) capture. In plants, these incongruences are most often seen between data derived from the nuclear vs. the cp genomes and the nuclear markers used for comparison usually have been from the nuclear ribosomal (nr) internal transcribed spacer region (ITS). The amount of genomic material shared between introgressing species can be highly variable. In some of these cases, other nuclear genomic regions have moved between species without leaving a signature on the nrITS. An example of well-supported phylogenetic incongruence is the placement of Dubautia scabra (DC.) D. D. Keck in the Hawaiian silversword alliance (HSA); evolutionary hypotheses for D. scabra based on molecular as opposed to cytogenetic data are strongly discordant. In this paper, we test these two conflicting phylogenetic hypotheses regarding the evolutionary relationships of Dubautia scabra using evidence from six low-copy nuclear genes, as well as multiple chloroplast noncoding regions and nrITS. The nrITS region is also examined for the presence of multiple copy types. Incongruence between inferred relationships based on nuclear chromosomal arrangements and molecular phylogenetic data from chloroplast DNA and nrITS is resolved in favor of a hypothesis of ancient hybridization rather than cytogenetic homoplasy involving dysploidy. Most single-copy nuclear genes track histories of D. scabra compatible with cytogenetic data whereas chloroplast and nrITS data track a common, different history that appears to reflect hybridization with a chromosomally distinct lineage that also occurs on Maui Nui and Hawai'i (the Big Island).

Sequence variation of four chloroplast non-coding regions among wild, weedy and cultivated Vigna angularis accessions

Sequence variation of four chloroplast non-coding regions among wild, weedy and cultivated Vigna angularis accessions

The architecture of the chloroplast psbA-trnH non-coding region in angiosperms

The psbA-trnH intergenic region is among the most variable regions in the angiosperm chloroplast genome. It is a popular tool for plant population genetics and species level phylogenetics and has been proposed as suitable for DNA barcoding studies. This region contains two parts differing in their evolutionary conservation: 1) the psbA 3′UTR (untranslated region) and 2) the psbA-trnH intergenic non-transcribed spacer. We compared the sequence and RNA secondary structure of the psbA 3′ UTR across angiosperms and found consensus motifs corresponding to the stem portions of the RNA stem-loop structures and a consensus TTAGTGTATA box. The psbA-trnH spacer exhibited patterns that can be explained by the independent evolution of large inversions in the psbA 3′UTR and mutational hot spots in the remaining portion of the psbA-trnH spacer. We conclude that a comparison of chloroplast UTRs across angiosperms offer clues to the identity of putative regulatory elements and information about selective constraints imposed on the chloroplast non-coding regions.

The molecular phylogeny of Rebutia (Cactaceae) and its allies demonstrates the influence of paleogeography on the evolution of South American mountain cacti

The tropical Andes harbor a major part of the world's plant biodiversity. The montane cacti of the tribes Browningieae, Cereeae, and Trichocereeae underwent extensive radiation and thus are well suited as a model group to study the diversification of Andean plants. We reconstructed their phylogeny employing three noncoding chloroplast regions and explained it in the context of the geological history of South America. We found that the clade of cephalia-bearing cacti with naked pericarpels is centered in northeastern Brazil, whereas almost all other clades comprise Andean species. The spatial split between the clades was probably caused by the Andean uplift and the concurrent formation of intracontinental marine basins in the Tertiary. The phylogenetic reconstructions based on parsimony and Bayesian approaches do not reflect the traditional delimitation of the tribes and of the large genera. Our results suggest that Rebutia s.l. and Echinopsis s.l. are not monophyletic and that Sulcorebutia, Weingartia, and Cintia should be united into one genus. Even though this "Weingartia-complex" and the genus Gymnocalycium are similar in size and morphological diversity, Gymnocalycium has a very high molecular divergence suggesting a comparably older radiation.

Phylogeny and biogeography of Sassafras (Lauraceae) disjunct between eastern Asia and eastern North America

Sassafras (Lauraceae) consists of three species disjunct between eastern Asia (S. tzumu and S. randaiense) and eastern North America (S. albidum). Phylogenetic analysis based on sequences of nuclear ribosomal ITS and three chloroplast non-coding regions (rpl16, trnL-F, and psbA-trnH) showed that Sassafras is monophyletic and that the eastern North American S. albidum is sister to the clade of its two eastern Asian counterparts. Their intercontinental divergence was estimated to be 13.80 ± 2.29−16.69 ± 2.52 million years ago (mya) using the penalized likelihood method with the ITS and three chloroplast markers. Biogeographic analyses combined with fossil evidence suggest that Sassafras has a relict distribution in the Northern Hemisphere without a Gondwanan link. The divergence time of the two eastern Asian species (the continental Chinese Sassafras tzumu and S. randaiense endemic to Taiwan) is estimated to be 0.61 ± 0.75−2.23 ± 0.76 mya. Sassafras randaiense from Taiwan was most likely derived from an ancestor from continental China.

Identification of Aegilops L. species and Triticum aestivum L. based on chloroplast DNA

The genus Aegilops L. is a very important genetic resource for the breeding of bread wheat Triticum aestivum. Therefore, an accurate and easy identification of Aegilops species is required. Traditionally, identification of Aegilops species has relied heavily on morphological characters. These characters, however, are either not variable enough among Aegilops species or too plastic to be used for identification at the species level. Molecular markers that are more stable within species, therefore, could be the alternative strategy towards an accurate identification. Since the chloroplast DNA has a lower level of evolution compared to the nuclear genome, an attempt was made in this study to investigate polymorphism in the chloroplast DNA among 21 Aegilops species (including Ae. mutica that is now known as Amblyopyrum muticum) and between the latter and T. aestivum to generate markers for the diagnosis of all targeted species. Cleaved amplified polymorphic sequence (CAPS) applied on 22 coding and non-coding chloroplast regions using 80 endonucleases and sequencing of two of those regions revealed little polymorphism between T. aestivum and the various Aegilops species examined and to a less extent was the variation among Aegilops species. Polymorphism observed among species analysed allowed the discrimination of T. aestivum and 12 Aegilops species.

Spore Morphology in Relation to Phylogeny in the Fern Genus Elaphoglossum (Dryopteridaceae)

The perispore structure of Elaphoglossum was studied using a scanning electron microscope. Of the species examined, 119 corresponded to those used in a previously published phylogenetic analysis of the genus based on two chloroplast noncoding DNA regions, trnL‐trnF and rps4‐trnS. The spores of 102 additional species were examined for comparative purposes. Five perispore characters were scored for each species and optimized onto the previously published molecular tree. The morphology of the perispore and its character state changes are described and discussed in a phylogenetic context. Synapomorphies for major clades within the genus were identified, such as spines for the Neotropical species of sect. Squamipedia and perforations, spines, and cristae for subsect. Pachyglossa and a large subclade within sect. Setosa. This study is the largest done on perispore morphology in relation to phylogeny in a genus of ferns. Spore images of all species studied are available at http://www.plantsystematics.org.

Comparison of whole chloroplast genome sequences to choose noncoding regions for phylogenetic studies in angiosperms: the tortoise and the hare III

Although the chloroplast genome contains many noncoding regions, relatively few have been exploited for interspecific phylogenetic and intraspecific phylogeographic studies. In our recent evaluation of the phylogenetic utility of 21 noncoding chloroplast regions, we found the most widely used noncoding regions are among the least variable, but the more variable regions have rarely been employed. That study led us to conclude that there may be unexplored regions of the chloroplast genome that have even higher relative levels of variability. To explore the potential variability of previously unexplored regions, we compared three pairs of single-copy chloroplast genome sequences in three disparate angiosperm lineages: Atropa vs. Nicotiana (asterids); Lotus vs. Medicago (rosids); and Saccharum vs. Oryza (monocots). These three separate sequence alignments highlighted 13 mutational hotspots that may be more variable than the best regions of our former study. These 13 regions were then selected for a more detailed analysis. Here we show that nine of these newly explored regions (rpl32-trnL((UAG)), trnQ((UUG))-5'rps16, 3'trnV((UAC))-ndhC, ndhF-rpl32, psbD-trnT((GGU)), psbJ-petA, 3'rps16-5'trnK((UUU)), atpI-atpH, and petL-psbE) offer levels of variation better than the best regions identified in our earlier study and are therefore likely to be the best choices for molecular studies at low taxonomic levels.

PHYLOGENETICS OF ACER (ACEROIDEAE, SAPINDACEAE) BASED ON NUCLEOTIDE SEQUENCES OF TWO CHLOROPLAST NON-CODING REGIONS

ABSTRACT Acer is one of the most diverse woody genera in the Northern Hemisphere. Recent phylogenetic studies support the placement of Acer and Dipteronia—sole members of the traditional Aceraceae—in the Sapindaceae. However, the monophyly of Acer and its sections remain to be tested. In this study, sequences of two chloroplast non-coding regions, psbM-trnD and trnD-trnT, are used to elucidate phylogenetic relationships of Acer and Dipteronia. Our results support the monophyly of Acer and sects. Arguta, Ginnala, Integrifolia, Lithocarpa, Macrantha, Palmata, Platanoidea, and Trifoliata. In contrast, sects. Acer, Goniocarpa, Parviflora, Saccharodendron, and Spicata are not monophyletic. Acer trautvetteri and A. opalus of sect. Acer are more closely related to A. monspessulanum of sect. Goniocarpa and A. saccharum of sect. Saccharina than to A. caesium and A. pseudoplatanus of sect. Acer. Acer distylium of sect. Parviflora is more closely related to sect. Platanoidea than to A. nipponicum of sect. Parviflora...

Molecular Markers for the Classification of Switchgrass (Panicum virgatum L.) Germplasm and to Assess Genetic Diversity in Three Synthetic Switchgrass Populations

Information regarding the amount of genetic diversity is necessary to enhance the effectiveness of breeding programs and germplasm conservation efforts. Genetic variation between 21 switchgrass genotypes randomly selected from two lowland (‘Alamo’ and ‘Kanlow’) and one upland (‘Summer’) synthetic cultivars were estimated using restriction fragment length polymorphism (RFLP) markers. Comparison of 85 RFLP loci revealed 92% polymorphism between at least two genotypes from the upland and lowland ecotypes. Within ecotypes, the upland genotypes showed higher polymorphism than lowland genotypes (64% vs. 56%). ‘Kanlow’ had a lower percent of polymorphic loci than ‘Alamo’ (52% vs. 60%). Jaccard distances revealed higher genetic diversity between upland and lowland ecotypes than between genotypes within each ecotype. Hierarchical cluster analysis using Ward's minimum variance grouped the genotypes into two major clusters, one representing the upland group and the other the lowland group. Phylogenetic analysis of chloroplast non-coding region trnL (UAA) intron sequences from 34 switchgrass accessions (6 upland cultivars, 2 lowland cultivars, and 26 accessions of unknown affiliation) produced a neighbor-joining dendrogram comprised of two major clusters with 99% bootstrap support. All accessions grouped in the same cluster with the lowland cultivars (‘Alamo’ and ‘Kanlow’) had a deletion of 49 nucleotides. Phenotypic identification of greenhouse-grown plants showed that all accessions with the deletion are of the lowland type. The deletion in trnL (UAA) sequences appears to be specific to lowland accessions and should be useful as a DNA marker for the classification of upland and lowland germplasm.

Phylogeny and biogeography of Altingiaceae: Evidence from combined analysis of five non-coding chloroplast regions

The Altingiaceae consist of ∼15 species that are disjunctly distributed in Asia and North America. The genus Liquidambar has been employed as a biogeographic model for studying the Northern Hemisphere intercontinental disjunctions. Parsimony and Bayesian analyses based on five non-coding chloroplast regions support that (1) Liquidambar is paraphyletic; (2) the temperate Liquidambar acalycina and Liquidambar formosana are nested within a large tropical to subtropical Asian clade; (3) Semiliquidambar is scattered in the eastern Asian clade and is of hybrid origin involving at least two maternal species: L. formosana and L. acalycina; and (4) the eastern North American Liquidambar styraciflua groups with the western Asian Liquidambar orientalis, but is highly distinct from other lineages. Biogeographically, our results demonstrate the complexity of biogeographic migrations throughout the history of Altingiaceae since the Cretaceous, with migration across both the Bering and the North Atlantic land bridges.

<I>Leavenworthia</I> (Brassicaceae) Revisited: Testing Classic Systematic and Mating System Hypotheses

The genus Leavenworthia (Brassicaceae) has long been a focus of research on mating system evolution, due to the presence of both self-incompatible and self-compatible species within the genus. A phylogenetic hypothesis invoking multiple transitions between mating systems has been proposed for Leavenworthia, but this hypothesis has not been subject to molecular phylogenetic analysis. DNA sequence variation from four non-coding chloroplast regions (the trnL intron; and the trnT-trnL, trnL-trnF, and psbA-trnH intergenic spacers) was used to reconstruct the generic phylogeny, to test the validity of several proposed species, and to assess the number of mating system transitions within the genus. The strict consensus tree largely reflected the long-standing phylogenetic hypothesis for Leavenworthia, although additional data are needed to fully validate the recognition of L. crassa and L. alabamica. Unexpected results included the placement of L. uniflora as sister to the rest of the genus, and the apparent hybridization between L. exigua and L. torulosa. Finally, our data strongly supported a minimum of three mating system transitions within Leavenworthia.