Gentianales Research Articles

Cytotoxic and anti-metastatic action mediates the anti-proliferative activity of Rhazya stricta Decne inducing apoptotic cell death in human cancer cells: Implication in chemopreventive mechanism

Cancer is associated with a high rate of pre-mature mortality worldwide and serves as a significant impediment to extended life expectancy, leading to undesirable long-term effects on the quality of life of the patients. Bioactive chemical products derived from natural sources have an important place in the well-being of the human population since ancient times. Compounds derived from natural sources have dominated the drug discovery programs in the last five decades and comprise a substantial proportion of current-day pharmaceutical agents. Rhazya stricta, an evergreen shrub, it belongs to family Apocynaceae of order Gentianales, and the Rhazya stricta is known to have certain medicinal properties, as the Middle East and South Asian traditional medicine employed this plant as a cure for different diseases. Herein, the hydro-methanolic extract of Rhazya stricta Decne has been studied for its effect on spectrum of cancer cells, including LNCaP, C4-2B, PC-3 prostate cancer cells, MDA-MB-231, MCF-7, SKBR3 breast cancer cells, A-549 lung cancer cells and BxPC-3, MiaPaCa-2 pancreatic cancer. Hydro-methanolic extract of Rhazya stricta Decne in a dose-dependent manner, showed progressive inhibition of cell growth by induction of cell death in the treated human cancer cells. The IC value range for different breast cancer cell lines was 142-178 µg/ml; for prostate cancer cell lines, it was 90-148 µg/ml; for pancreatic cancer cell lines 116-166 µg/ml and for the lung cancer cells, it was reported to be 180 µg/ml. The anti-metastatic potential of extract was indicated by the notable reduction in invasiveness and cell migration. Moreover, the structurally diverse phytochemical rich extract, also downregulates the signaling of NF-kB and the NF-kB downstream target cytokine VEGF, providing an insight into the anticancer action mechanism.Keywords: Cell biology; Cancer chemoprevention; Anticancer drugs; Cell signaling; Natural compounds

Traditional Healer of Asclepiadoideae of Apocynaceae in Bankura, West Bengal

The present communication deals with the ethno-botanical exploration, identification of some plants of formerly Asclepiadaceae that are treated as subfamily Asclepiadoideae, the milkweed subfamily of family Apocynaceae (order Gentianales) as per APG III system of classification. The plants of this subfamily are being used frequently by the rural people of Bankura. During the present investigation, a total of 14 species of subfamily Asclepiadoideae have been reportedthey were used by the local health healers for the treatment of different diseases. Among them Calotropisgigantea, Hemidesmusindicus, Pergulariadaemia, Telosmapallida, Tylophoraindica, are leading species that are frequently used for a variety of health problems. As reported by informants, roots are the most important part used for herbal preparation followed by whole plant, leaf and fruit. The rural people of Bankura have good knowledge about ethno-medicinal plants and this legacy of traditional culture must be conserved. So present paper depicts various plant resources of ethno-medicinewith special reference to subfamily Asclepiadoideae.

A complete sequence of mitochondrial genome of Neolamarckia cadamba and its use for systematic analysis

Neolamarckia cadamba is an important tropical and subtropical tree for timber industry in southern China and is also a medicinal plant because of the secondary product cadambine. N. cadamba belongs to Rubiaceae family and its taxonomic relationships with other species are not fully evaluated based on genome sequences. Here, we report the complete sequences of mitochondrial genome of N. cadamba, which is 414,980 bp in length and successfully assembled in two genome circles (109,836 bp and 305,144 bp). The mtDNA harbors 83 genes in total, including 40 protein-coding genes (PCGs), 31 transfer RNA genes, 6 ribosomal RNA genes, and 6 other genes. The base composition of the whole genome is estimated as 27.26% for base A, 22.63% for C, 22.53% for G, and 27.56% for T, with the A + T content of 54.82% (54.45% in the small circle and 54.79% in the large circle). Repetitive sequences account for ~ 0.14% of the whole genome. A maximum likelihood (ML) tree based on DNA sequences of 24 PCGs supports that N. cadamba belongs to order Gentianales. A ML tree based on rps3 gene of 60 species in family Rubiaceae shows that N. cadamba is more related to Cephalanthus accidentalis and Hymenodictyon parvifolium and belongs to the Cinchonoideae subfamily. The result indicates that N. cadamba is genetically distant from the species and genera of Rubiaceae in systematic position. As the first sequence of mitochondrial genome of N. cadamba, it will provide a useful resource to investigate genetic variation and develop molecular markers for genetic breeding in the future.

The complete chloroplast genome of alien invasive species, Diodia virginiana L. (rubiaceae), in China

The first complete chloroplast genome sequences of new intercepted alien weeds in China, Diodia, were reported in this study. The D. virginiana plastome was 154,387 bp long, with the large single copy (LSC) region of 83,625 bp, the small single copy (SSC) region of 17,475 bp, and two inverted repeat (IR) regions of 26,643 bp. The plastome contained 137 genes, including 92 proteincoding, eight ribosomal RNA, and 37 transfer RNA genes. The overall GC content was 37.2%. Phylogenetic analysis of 24 representative plastomes within the order Gentianales suggests that subfamily Cinchonoideae is closely related to subfamily Ixoroideae compared with subfamily Rubioideae in Rubiaceae. Tribe Spermacoceae was most closely related to Rubieae among the three representative groups of subfamily Rubioideae.

Cytochrome P450 and O-methyltransferase catalyze the final steps in the biosynthesis of the anti-addictive alkaloid ibogaine from Tabernanthe iboga

Monoterpenoid indole alkaloids are a large (∼3000 members) and structurally diverse class of metabolites restricted to a limited number of plant families in the order Gentianales. Tabernanthe iboga or iboga (Apocynaceae) is native to western equatorial Africa and has been used in traditional medicine for centuries. Howard Lotsof is credited with bringing iboga to the attention of Western medicine through his accidental discovery that iboga can alleviate opioid withdrawal symptoms. Since this observation, iboga has been investigated for its use in the general management of addiction. We were interested in elucidating ibogaine biosynthesis to understand the unique reaction steps en route to ibogaine. Furthermore, because ibogaine is currently sourced from plant material, these studies may help improve the ibogaine supply chain through synthetic biology approaches. Here, we used next-generation sequencing to generate the first iboga transcriptome and leveraged homology-guided gene discovery to identify the penultimate hydroxylase and final O-methyltransferase steps in ibogaine biosynthesis, herein named ibogamine 10-hydroxylase (I10H) and noribogaine-10-O-methyltransferase (N10OMT). Heterologous expression in Saccharomyces cerevisiae (I10H) or Escherichia coli (N10OMT) and incubation with putative precursors, along with HPLC–MS analysis, confirmed the predicted activities of both enzymes. Moreover, high expression levels of their transcripts were detected in ibogaine-accumulating plant tissues. These discoveries coupled with our publicly available iboga transcriptome will contribute to additional gene discovery efforts and could lead to the stabilization of the global ibogaine supply chain and to the development of ibogaine as a treatment for addiction.

Blind Testing: DNA Barcoding Sheds Light Upon the Identity of Plant Fragments as a Subsidy for Cave Conservation

Plants living above and around caves represent an important, albeit poorly studied, resource within cave ecosystems. The presence of plant material (root-like structures or rhizothemes, saplings, seeds, and seedlings) correlates positively with the biodiversity of the cave dwelling animals as shown for iron-ore caves in Carajás, Pará, Brazil. Plant material collected in caves has proven to be difficult to identify by traditional botanical methods, thus this research aims to provide a qualitative insight into the taxonomy and morphology of rhizothemes and other plant fragments found in the caves. The identification process used a combination of different molecular markers (ITS2, rbcL, and trnH-psbA) followed by a comparison of the sequences obtained against publicly available databases. The rhizothemes were submitted to micromorphological analysis to ascertain their putative root or stem origin and to compare their anatomy with known patterns found in the plant families or genera recovered through molecular matches. All studied samples were Angiosperms, mostly belonging to subclass Rosideae, within four orders: Malpighiales (Euphorbiaceae, Hypericaceae), Sapindales (Anacardiaceae and Sapindaceae), Myrtales (Myrtaceae), Fabales (Fabaceae), and only two belonging to subclass Asteridae, order Gentianales (Apocynaceae). Some of the samples were matched to generic level, with ITS2 being the best marker to identify the fragments because it shows high degree of sequence variation even at specific level and result reliability. All rhizothemes turned out to be roots, and correspondence was found between the existing literature and the individual anatomical patterns for the families and genera retrieved. DNA barcode has proved to be a useful tool to identify plant fragments found in this challenging environment. However, the existence of well curated, authoritatively named collections with ample biological information has proven to be essential to achieve a reliable identification.

Explaining Ovule Sterility inRauvolfiaL. (Apocynaceae)

Premise of research. Approximately 5% of Gentianales are estimated to be dioecious—mainly the members of Rubiaceae, Gentianaceae, and Loganiaceae. In Apocynaceae, there exists a report of cryptic d...

Resolution effects in reconstructing ancestral genomes

BackgroundThe reconstruction of ancestral genomes must deal with the problem of resolution, necessarily involving a trade-off between trying to identify genomic details and being overwhelmed by noise at higher resolutions.ResultsWe use the median reconstruction at the synteny block level, of the ancestral genome of the order Gentianales, based on coffee, Rhazya stricta and grape, to exemplify the effects of resolution (granularity) on comparative genomic analyses.ConclusionsWe show how decreased resolution blurs the differences between evolving genomes, with respect to rate, mutational process and other characteristics.

Chloroplast genome structures in Gentiana (Gentianaceae), based on three medicinal alpine plants used in Tibetan herbal medicine.

The genus Gentiana is the largest in the Gentianaceae family with ca. 400 species. However, with most species growing on the Qinghai-Tibet plateau, the processes of adaptive evolution and speciation within the genus is not clear. Also, the genomic analyses could provide important information. So far, the complete chloroplast (cp) genome data of the genus are still deficient. As the second and third sequenced members within Gentianaceae, we report the construction of complete cp sequences of Gentiana robusta King ex Hook. f. and Gentiana crassicaulis Duthie ex Burk., and describe a comparative study of three Gentiana cp genomes, including the cp genome of Gentiana straminea Maxim. published previously. These cp genomes are highly conserved in gene size, gene content, and gene order and the rps16 pseudogene with exon2 missing was found common. Three repeat types and five SSR types were investigated, and the number and distribution are similar among the three genomes. Sixteen genome divergent hotspot regions were identified across these cp genomes that could provide potential molecular markers for further phylogenetic studies in Gentiana. The IR/SC boundary organizations in Gentianales cp genomes were compared and three different types of boundaries were observed. Six data partitions of cp genomes in Gentianales were used for phylogenetic analyses and different data partitions were largely congruent with each other. The ML phylogenetic tree was constructed based on the fragments in cp genomes commonly available in 33 species from Lamiids, including 12 species in Gentianales, 1 in Boraginaceae, 10 in Solanales, and 10 in Lamiales. The result strongly supports the position of Boraginaceae (Ehretia acuminata) as the sister of Solanales, with the bootstrap values of 97%. This study provides a platform for further research into the molecular phylogenetics of species in the order Gentianales (family Gentianaceae) notably in respect of speciation and species identification.

The complete chloroplast genome of Gentiana straminea (Gentianaceae), an endemic species to the Sino-Himalayan subregion

Endemic to the Sino-Himalayan subregion, the medicinal alpine plant Gentiana straminea is a threatened species. The genetic and molecular data about it is deficient. Here we report the complete chloroplast (cp) genome sequence of G. straminea, as the first sequenced member of the family Gentianaceae. The cp genome is 148,991bp in length, including a large single copy (LSC) region of 81,240bp, a small single copy (SSC) region of 17,085bp and a pair of inverted repeats (IRs) of 25,333bp. It contains 112 unique genes, including 78 protein-coding genes, 30 tRNAs and 4 rRNAs. The rps16 gene lacks exon2 between trnK-UUU and trnQ-UUG, which is the first rps16 pseudogene found in the nonparasitic plants of Asterids clade. Sequence analysis revealed the presence of 13 forward repeats, 13 palindrome repeats and 39 simple sequence repeats (SSRs). An entire cp genome comparison study of G. straminea and four other species in Gentianales was carried out. Phylogenetic analyses using maximum likelihood (ML) and maximum parsimony (MP) were performed based on 69 protein-coding genes from 36 species of Asterids. The results strongly supported the position of Gentianaceae as one member of the order Gentianales. The complete chloroplast genome sequence will provide intragenic information for its conservation and contribute to research on the genetic and phylogenetic analyses of Gentianales and Asterids.

Identification of novel and conserved microRNAs in Coffea canephora and Coffea arabica.

As microRNAs (miRNAs) are important regulators of many biological processes, a series of small RNAomes from plants have been produced in the last decade. However, miRNA data from several groups of plants are still lacking, including some economically important crops. Here microRNAs from Coffea canephora leaves were profiled and 58 unique sequences belonging to 33 families were found, including two novel microRNAs that have never been described before in plants. Some of the microRNA sequences were also identified in Coffea arabica that, together with C. canephora, correspond to the two major sources of coffee production in the world. The targets of almost all miRNAs were also predicted on coffee expressed sequences. This is the first report of novel miRNAs in the genus Coffea, and also the first in the plant order Gentianales. The data obtained establishes the basis for the understanding of the complex miRNA-target network on those two important crops.

Gelsemiaceae (Gentianales) expanded to include the enigmatic Asian genusPteleocarpa

The enigmatic South-East Asian monotypic genus Pteleocarpa has been considered as a genus incertae sedis among the eudicots for a long time. Molecular data (plastid and nuclear ribosomal regions) from 44 widely sampled species across Lamiidae and phylogenetic analyses have finally clarified its familial relationships, and it is here included in Gelsemiaceae (order Gentianales). Its morphological characteristics support a placement in this family and order as a result of the presence of potential synapomorphies, such as imbricate and commonly yellow corollas, latrorse anther dehiscence, divided styles and compressed seeds. Unique characters for Pteleocarpa in Gelsemiaceae are alternate leaves and indehiscent samaras. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 175, 482–496.

Cyclotide discovery in Gentianales revisited--identification and characterization of cyclic cystine-knot peptides and their phylogenetic distribution in Rubiaceae plants.

Cyclotides are a unique class of ribosomally synthesized cysteine-rich miniproteins characterized by a head-to-tail cyclized backbone and three conserved disulfide-bonds in a knotted arrangement. Originally they were discovered in the coffee-family plant Oldenlandia affinis (Rubiaceae) and have since been identified in several species of the violet, cucurbit, pea, potato, and grass families. However, the identification of novel cyclotide-containing plant species still is a major challenge due to the lack of a rapid and accurate analytical workflow in particular for large sampling numbers. As a consequence, their phylogeny in the plant kingdom remains unclear. To gain further insight into the distribution and evolution of plant cyclotides, we analyzed ∼300 species of >40 different families, with special emphasis on plants from the order Gentianales. For this purpose, we have developed a refined screening methodology combining chemical analysis of plant extracts and bioinformatic analysis of transcript databases. Using mass spectrometry and transcriptome-mining, we identified nine novel cyclotide-containing species and their related cyclotide precursor genes in the tribe Palicoureeae. The characterization of novel peptide sequences underlines the high variability and plasticity of the cyclotide framework, and a comparison of novel precursor proteins from Carapichea ipecacuanha illustrated their typical cyclotide gene architectures. Phylogenetic analysis of their distribution within the Psychotria alliance revealed cyclotides to be restricted to Palicourea, Margaritopsis, Notopleura, Carapichea, Chassalia, and Geophila. In line with previous reports, our findings confirm cyclotides to be one of the largest peptide families within the plant kingdom and suggest that their total number may exceed tens of thousands.

The Complete Plastid Genome Sequence of Madagascar Periwinkle Catharanthus roseus (L.) G. Don: Plastid Genome Evolution, Molecular Marker Identification, and Phylogenetic Implications in Asterids.

The Madagascar periwinkle ( Catharanthus roseus in the family Apocynaceae) is an important medicinal plant and is the source of several widely marketed chemotherapeutic drugs. It is also commonly grown for its ornamental values and, due to ease of infection and distinctiveness of symptoms, is often used as the host for studies on phytoplasmas, an important group of uncultivated plant pathogens. To gain insights into the characteristics of apocynaceous plastid genomes (plastomes), we used a reference-assisted approach to assemble the complete plastome of C . roseus , which could be applied to other C . roseus -related studies. The C . roseus plastome is the second completely sequenced plastome in the asterid order Gentianales. We performed comparative analyses with two other representative sequences in the same order, including the complete plastome of Coffea arabica (from the basal Gentianales family Rubiaceae) and the nearly complete plastome of Asclepias syriaca (Apocynaceae). The results demonstrated considerable variations in gene content and plastome organization within Apocynaceae, including the presence/absence of three essential genes (i.e., accD, clpP, and ycf1) and large size changes in non-coding regions (e.g., rps2-rpoC2 and IRb-ndhF). To find plastome markers of potential utility for Catharanthus breeding and phylogenetic analyses, we identified 41 C . roseus -specific simple sequence repeats. Furthermore, five intergenic regions with high divergence between C . roseus and three other euasterids I taxa were identified as candidate markers. To resolve the euasterids I interordinal relationships, 82 plastome genes were used for phylogenetic inference. With the addition of representatives from Apocynaceae and sampling of most other asterid orders, a sister relationship between Gentianales and Solanales is supported.

Patterns of Embryological and Biochemical Evolution in the Asterids

Abstract The Monopetalae or Sympetalae have been resurrected recently as the Asteridae s.l., a large clade that includes almost all sympetalous angiosperms. Phylogenetic analyses of a large, combined data set of sequences of four genes (18S rDNA, rbcL, ndhF, atpB) have improved our understanding of the evolutionary history of this large clade of angiosperms. Most of the taxa fall within one of four major clades, the Cornales, Ericales, euasterids I (comprising Lamiales, Solanales, Boraginaceae, Gentianales, and Garryales), and euasterids II (comprising Asterales, Apiales, Dipsacales, Escalloniales, and Aquifoliales). We used the phylogenetic tree based on our recent phylogenetic analysis of four genes to elucidate the evolutionary patterns of the ovule and the diversification of iridoid biosynthesis in the asterids. Asterid-specific patterns of character distribution can be seen. Unitegmic-tenuinucellate ovules occur almost universally in the asterid clade. Cellular endosperm is ancestral in the asterids;...

Chemical constiuents from Richardia grandiflora (Cham. & Schltdl.) Steud. (Rubiaceae)

Amongst the different forms of therapy to prevent and cure illnesses, plants have been, undoubtedly, the most utilized ones since the beginning of mankind. Brazil has a great diversity on plants that possess non-researched medicinal potential and are promising sources of therapeutic and pharmacological innovations. The Rubiaceae family is considered the biggest one of the order Gentianales, presenting around 637 genera and 10,700 species. Richardia grandiflora (Cham. & Schltdl.) Steud., known popularly as "ervanço", "poaia" or "ipeca-mirim", has ethnopharmacological indications to use as decoction against hemorrhoids and as vermifuge. Aiming at contributing to the chemotaxonomic study of the family Rubiaceae and considering the absence of data in literature about the chemical constitution of the species Richardia grandiflora, the latter was submitted to a phytochemical study to isolate its chemical constituents, through usual chromatographic methods, and after identifying them by means of spectroscopic methods such as ¹H and 13C NMR, with the add of two-dimensional techniques, besides comparison with literature data. Five constituents were isolated through this first phytochemical study with R. grandiflora: a mixture of the steroids beta-sitosterol and stigmasterol, o-hydroxy-benzoic acid, m-methoxy-p-hydroxy-benzoic acid and phaeophitin A, all of them isolated for the first time from the genus Richardia.

Nectar sugar composition and volumes of 47 species of Gentianales from a southern Ecuadorian montane forest.

This study investigates 47 taxonomically related species (Gentianales), all native to a tropical montane forest in southern Ecuador, in terms of nectar chemistry and nectar volumes in relation to pollination biology. Nectar volumes of covered (24-h production) and uncovered (standing crop) flowers were measured in the natural habitat. Sucrose, fructose and glucose were quantified in the nectar using high performance liquid chromatography. Flower visitors were observed. Nectar sugar concentration did not differ significantly among the pollination syndromes. Regarding sugar composition, the only significant differences were found in chiropterophilous and myiophilous flowers, which had a significantly lower sugar ratio than sphingophilous flowers. A separation of chiropterophilous and myiophilous flowers from the other pollination syndromes is further substantiated by non-linear multidimensional scaling using the chord-normalized expected species shared index of dissimilarity based on nectar sugar compositions. The matrix test revealed no correlation of observed floral visitors to nectar concentrations; however, a weak significant correlation was found between floral visitors and nectar sugar compositions. The nectar volumes of covered and uncovered flowers are related to, and differ significantly among, pollination syndromes. Matrix tests revealed correlation between floral visitors and nectar volume of covered flowers and, to a lesser extent, of uncovered flowers. Sucrose is the predominant floral nectar sugar in the order Gentianales, suggesting that nectar sugar composition is a conservative characteristic. However, some degree of an adaptive convergence of floral nectar compositions to principal pollinator type within the constraints set by phylogenetic history is likely. The driving force to visitation appears to be the volume of nectar the visitor can expect to consume.

Structural analysis of xyloglucans in the primary cell walls of plants in the subclass Asteridae

The structures of xyloglucans from several plants in the subclass Asteridae were examined to determine how their structures vary in different taxonomic orders. Xyloglucans, solubilized from plant cell walls by a sequential (enzymatic and chemical) extraction procedure, were isolated, and their structures were characterized by NMR spectroscopy and mass spectrometry. All campanulids examined, including Lactuca sativa (lettuce, order Asterales), Tenacetum ptarmiciflorum (dusty miller, order Asterales), and Daucus carota (carrot, order Apiales), produce typical xyloglucans that have an XXXG-type branching pattern and contain α- d-Xyl p-, β- d-Gal p-(1→2)-α- d-Xyl p-, and α- l-Fuc p-(1→2)-β- d-Gal p-(1→2)-α- d-Xyl p- side chains. However, the lamiids produce atypical xyloglucans. For example, previous analyses showed that Capsicum annum (pepper) and Lycopersicon esculentum (tomato), two species in the order Solanales, and Olea europaea (olive, order Lamiales) produce xyloglucans that contain arabinosyl and galactosyl residues, but lack fucosyl residues. The XXGG-type xyloglucans produced by Solanaceous species are less branched than the XXXG-type xyloglucan produced by Olea europaea. This study shows that Ipomoea pupurea (morning glory, order Solanales), Ocimum basilicum (basil, order Lamiales), and Plantago major (plantain, order Lamiales) all produce xyloglucans that lack fucosyl residues and have an unusual XXGGG-type branching pattern in which the basic repeating core contains five glucose subunits in the backbone. Furthermore, Neruim oleander (order Gentianales) produces an XXXG-type xyloglucan that contains arabinosyl, galactosyl, and fucosyl residues. The appearance of this intermediate xyloglucan structure in oleander has implications regarding the evolutionary development of xyloglucan structure and its role in primary plant cell walls.

The IBOGA alkaloids and their role as precursors of anti-neoplastic bisindole catharanthus alkaloids

Iboga alkaloids are found in plants of the family Apocynaceae, which falls within the order Gentianales. The iboga alkaloids are examples of the broad class of monoterpenoid indole alkaloids that are considered a chemical marker for the Apocynaceae. This chapter discusses the chemistry of iboga alkaloids and their role as precursors of anti-neoplastic bisindole Catharanthus alkaloids. The iboga alkaloids are comprised of indole and isoquinuclidine rings fused by a seven-membered C-ring. There is a carboxy group at C-16 in the C20 alkaloids, but other iboga alkaloids lack this substituent. Ibogamine and catharanthine are prototypical structures. Studies have shown the catharanthine isolated from natural sources is usually of the opposite absolute configuration from ibogamine. The chapter uses the biosynthetic numbering scheme of Le Men and Taylor. The chapter also discusses the structures in which the C-16-C-2 1 bond is cleaved. The C-16,C-21 fragmented structure occurs in the anti-neoplastic bisindole alkaloids isolated from Catharanthus roseus. The iboga structure also frequently occurs in combination with the vobasan skeleton.

Discovery of paralogous nuclear gene sequences coding for the second-largest subunit of RNA polymerase II (RPB2) and their phylogenetic utility in gentianales of the asterids.

Paralogous sequences of the RPB2 gene are demonstrated in the angiosperm order Gentianales. Two different copies were found by using different PCR primer pairs targeting a region that corresponds to exons 22-24 in the Arabidopsis RPB2 gene. One of the copies (RPB2-d) lacks introns in this region, whereas the other has introns at locations corresponding to those of green plants previously investigated. When analyzed with other available RPB2 sequences from this region, all 28 RPB2-d sequences obtained from the Gentianales and the four sequences from the Lamiales form a monophyletic group, together with a previously published tomato cDNA sequence. The substitution patterns, relative rates of change, and nucleotide compositions of the two paralogous RPB2 exon regions are similar, and none of them shows any signs of being a pseudogene. Although multiple copies of similar, paralogous sequences can confound phylogenetic interpretations, the lack of introns in RPB2-d make a priori homology assessment easy. The phylogenetic utility of RPB2-d within the Gentianales is evaluated in comparison with the chloroplast genes ndhF and rbcL. The hierarchical information in the RPB2-d region sequenced is more incongruent with that of the plastid genes than the plastid genes are with each other as determined by incongruence length difference tests. In contrast to the plastid genes, parsimony-informative third codon positions of RPB2 have a significantly higher rate of change than first and second positions. Topologically, the trees from the three genes are similar, and the differences are usually only weakly supported. In terms of support, RPB2 gives the highest jackknife support per sequenced nucleotide, whereas ndhF gives the highest Bremer support per sequenced nucleotide. The RPB2-d locus has the potential to be a valuable nuclear marker for determination of phylogenetic relationships within the euasterid I group of plants.