Chloroplast Fragments Research Articles

Transcriptome analysis of the Holly mangrove Acanthus ilicifolius and its terrestrial relative, Acanthus leucostachyus, provides insights into adaptation to intertidal zones.

BackgroundAcanthus is a unique genus consisting of both true mangrove and terrestrial species; thus, it represents an ideal system for studying the origin and adaptive evolution of mangrove plants to intertidal environments. However, little is known regarding the two respects of mangrove species in Acanthus. In this study, we sequenced the transcriptomes of the pooled roots and leaves tissues for a mangrove species, Acanthus ilicifolius, and its terrestrial congener, A. leucostachyus, to illustrate the origin of the mangrove species in this genus and their adaptive evolution to harsh habitats.ResultsWe obtained 73,039 and 69,580 contigs with N50 values of 741 and 1557 bp for A. ilicifolius and A. leucostachyus, respectively. Phylogenetic analyses based on four nuclear segments and three chloroplast fragments revealed that mangroves and terrestrial species in Acanthus fell into different clades, indicating a single origin of the mangrove species in Acanthus. Based on 6634 orthologs, A. ilicifolius and A. leucostachyus were found to be highly divergent, with a peak of synonymous substitution rate (Ks) distribution of 0.145 and an estimated divergence time of approximately 16.8 million years ago (MYA). The transgression in the Early to Middle Miocene may be the major reason for the entry of the mangrove lineage of Acanthus into intertidal environments. Gene ontology (GO) classifications of the full transcriptomes did not show any apparent differences between A. ilicifolius and A. leucostachyus, suggesting the absence of gene components specific to the mangrove transcriptomes. A total of 99 genes in A. ilicifolius were identified with signals of positive selection. Twenty-three of the 99 positively selected genes (PSGs) were found to be involved in salt, heat and ultraviolet stress tolerance, seed germination and embryo development under periodic inundation. These stress-tolerance related PSGs may be crucial for the adaptation of the mangrove species in this genus to stressful marine environments and may contribute to speciation in Acanthus.ConclusionsWe characterized the transcriptomes of one mangrove species of Acanthus, A. ilicifolius, and its terrestrial relative, A. leucostachyus, and provided insights into the origin of the mangrove Acanthus species and their adaptive evolution to abiotic stresses in intertidal environments.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1813-9) contains supplementary material, which is available to authorized users.

Influence of niche similarity on hybridization between Myriophyllum sibiricum and M.spicatum.

The impact of ecological factors on natural hybridization is of widespread interest. Here, we asked whether climate niche influences hybridization between the two closely related plant species Myriophyllum sibiricum and M.spicatum. Eight microsatellite loci and two chloroplast fragments were used to investigate the occurrence of hybridization between these two species in two co-occurring regions: north-east China (NEC) and the Qinghai-Tibetan Plateau (QTP). The climate niches of the species were quantified by principal component analysis with bioclimatic data, and niche comparisons were performed between the two species in each region. Reciprocal hybridization was observed, and M.sibiricum was favoured as the maternal species. Furthermore, hybrids were rare in NEC but common in the QTP. Accordingly, in NEC, the two species were climatically distinct, and hybrids only occurred in the narrow geographical or ecological transition zone, whereas in the QTP, obvious niche overlaps were found for the two species, and hybrids occurred in multiple contact zones. This association between hybridization pattern and climate niche similarity suggests that the level of hybridization was promoted by niche overlap. Compared with the parental species, similar climate niches were found for the hybrid populations in the QTP, indicating that other environmental factors rather than climate were important for hybrid persistence. Our findings highlight the significance of climate niche with respect to hybridization patterns in plants.

Multilocus species tree analyses resolve the ancient radiation of the subtribe Zizaniinae (Poaceae)

The phylogeny of the subtribe Zizaniinae of rice tribe (Oryzeae) has not been well resolved, particularly for the monotypic Hygroryza whose systematic position was inconsistent in previous studies. Here, we used the concatenation approach and coalescent-based species tree methods to reconstruct the phylogeny of Zizaniinae based on sequences of 14 nuclear single-copy loci and concatenated chloroplast fragments. Despite the low resolution of the tree from concatenated data and substantial topological incongruence of individual gene trees, the species trees inferred from three coalescent-based methods were fully concordant and highly supported. Importantly, the genus Hygroryza was consistently recovered with strong support by all coalescent-based methods. Further various phylogenetic analyses indicated that incomplete lineage sorting was the most likely process that generated pervasive discordance among individual gene trees, although hybridization and introgression cannot be excluded completely. Our species tree inferences based on multilocus data successfully resolved the phylogenetic relationships of the Zizaniinae lineages and confirmed that ancient rapid radiation has taken place in the diversification history of Zizaniinae. This study demonstrates that coalescent-based species tree approaches outperformed the concatenation method and could effectively decipher ancient rapid radiations as long as well resolved individual gene trees were sufficiently sampled.

Effects of domestication on genetic diversity in Chimonanthus praecox: Evidence from chloroplast DNA and amplified fragment length polymorphism data

AbstractChimonanthus praecox (L.) Link is a widely cultivated endemic winter‐flowering plant in China that has a long cultivation history. Genetic diversity and genetic structure were compared between wild and cultivated groups to reveal the geographic origin of the cultivated genotypes using chloroplast DNA (cpDNA) sequences and amplified fragment length polymorphism (AFLP) markers. Nine haplotypes were identified using three combined chloroplast fragments. Based on chloroplast data, the wild group showed greater genetic variation and genetic differentiation and a lower measure of gene flow compared to the cultivated group. The AFLP markers also supported this trend. More than 40% of the cpDNA haplotypes were shared between wild and cultivated groups, with shared haplotypes originating from multiple wild populations, suggesting multiple origins of cultivated plants. Moreover, principal coordinate analysis, UPGMA, and structure analysis of AFLP markers revealed that two wild populations clustered with most of the cultivated populations of Ch. praecox, suggesting that most of the cultivated populations mainly originated from these two populations. The combined cpDNA and AFLP results indicated that modern cultivated Ch. praecox experienced multiple events of origin involving two geographic origins, eastern China (Tianmu Mountain) and southwestern China (the border of Hunan–Guangxi–Sichuan–Guizhou).

Consistency between molecular phylogeny and morphological classification of the Salix matsudana Koidz. complex (Salicaceae).

The morphological species concept is based on morpho-logical traits, which are often subject to subjectivity or artifact. Molecular evidence is needed to test the reliability of morphological classification of taxa that are controversial and to provide appropriate taxonomic de-limitation. In this study, we used 15 single-copy nuclear loci and 2 chloroplast fragments to verify the morphological classification of the Salix matsudana Koidz. complex using phylogenetic approaches. Complete sequence alignment showed slight diversification in nuclear sequences and no variety in chloroplast DNA fragments. Phylogenetic trees revealed a monophyletic group consisting of all individuals of S. matsudana and 2 clades within this group, with a 100% bootstrap support value and 1.00 posterior probability. The topology of the phylogenetic trees was highly consistent with the morphological classification of the S. matsudana complex. Verifying the genetic background of these classification units based on remarkable morphological differences will provide a foundation for future studies of Salix and the breeding of new horticultural varieties.

Phylogeography of Rhodiola kirilowii (Crassulaceae): a story of Miocene divergence and quaternary expansion.

The evolution and current distribution of the Sino-Tibetan flora have been greatly affected by historical geological events, such as the uplift of the Qinghai-Tibetan Plateau (QTP), and Quaternary climatic oscillations. Rhodiola kirilowii, a perennial herb with its distribution ranging from the southeastern QTP and the Hengduan Mountains (HM) to adjacent northern China and central Asia, provides an excellent model to examine and disentangle the effect of both geological orogeny and climatic oscillation on the evolutionary history of species with such distribution patterns. We here conducted a phylogeographic study using sequences of two chloroplast fragments (trnL-F and trnS-G) and internal transcribed spacers in 29 populations of R. kirilowii. A total of 25 plastid haplotypes and 12 ITS ribotypes were found. Molecular clock estimation revealed deep divergence between the central Asian populations and other populations from the HM and northern China; this split occurred ca. 2.84 million year ago. The majority of populations from the mountains of northern China were dominated by a single haplotype or ribotype, while populations of the HM harbored both high genetic diversity and high haplotype diversity. This distribution pattern indicates that HM was either a diversification center or a refugium for R. kirilowii during the Quaternary climatic oscillations. The present distribution of this species on mountains in northern China may have resulted from a rapid glacial population expansion from the HM. This expansion was confirmed by the mismatch distribution analysis and negative Tajima's D and Fu's F S values, and was dated to ca. 168 thousand years ago. High genetic diversity and population differentiation in both plastid and ITS sequences were revealed; these imply restricted gene flow between populations. A distinct isolation-by-distance pattern was suggested by the Mantel test. Our results show that in old lineages, populations may harbour divergent genetic forms that are sufficient to maintain or even increase overall genetic diversity despite fragmentation and low within-population variation.

New insights into the hybrid origin of Malus toringoides and its close relatives based on a single‐copy nuclear gene SbeI and three chloroplast fragments

AbstractMalus toringoides Hughes and its close relatives, M. maerkangensis M. H. Cheng et al., M. setok Vassilcz., and M. xiaojinensis M. H. Cheng & N. G. Jiang were supposed to derive from hybridizations. However, molecular data are still inadequate to corroborate the hybrid origin hypotheses. In this study, we sequenced a single‐copy nuclear gene SbeI and three chloroplast fragments and carried out phylogenetic analyses to investigate the evolutionary origins of the above four putative hybrid taxa. The hybrid nature of M. toringoides is confirmed by the detection of two distinct types of SbeI sequences from it. The chloroplast and SbeI gene phylogenies show that the maternal progenitor of M. toringoides is closely related to M. sikkimensis N. P. Balakr. and M. spectabilis Borkh., and the paternal progenitor is most likely M. transitoria C. K. Schneid. The hypothesis that M. kansuensis (Batalin) C. K. Schneid. is one of the parents of M. toringoides is not supported. Malus maerkangensis and M. xiaojinensis might have originated through hybridization between M. toringoides and M. kansuensis, whereas M. setok is genetically closely related to M. toringoides. The three close relatives of M. toringoides were designated as three novel species by some researchers, however, as they were all apomictic with limited distribution areas and they originated from hybridization and polyploidization, we recommend that their species status should be re‐evaluated.

Genetic evidence for paternal inheritance of the chloroplast in four Australian Callitris species (Cupressaceae)

The mode of chloroplast inheritance was investigated in the four Australian conifer species; Callitris columellaris, C. intratropica, C. gracilis and C. verrucosa (Callitroideae, Cupressaceae), as a basis for future use of chloroplast genetic markers for the genus. Open-pollinated seeds and the foliage tissue of the mother trees were collected from one population per species, in which two distinct chloroplast haplotypes at the trnL (UAA) 5′ exon–trnF (GAA) region were co-distributed. In total, the DNA sequences of the chloroplast fragments in 255 samples were determined, and the haplotypes of the germinated seedlings and the mother trees were compared. Among the 16 mother trees examined across all species, 12 trees were shown to have produced at least one offspring with a different haplotype from the mother, indicating the occurrence of paternal inheritance of chloroplast in the fertilization processes. In binomial GLMM analyses, the haplotype of mother trees in C. columellaris, C. gracilis, and C. verrucosa did not significantly affect those of their offspring, and the predicted frequency of seedlings' haplotypes was compatible with the observed frequency in adult canopy trees. These findings provide a genetic support that paternal inheritance of chloroplast DNA via pollen dispersal would be dominant in the Callitris species.

Genetic structure of the Delphinium naviculare species group tracks Pleistocene climatic oscillations in the Tianshan Mountains, arid Central Asia

Pleistocene climatic fluctuations have affected the evolution of the genetic structure of many organisms. In this study, we test the hypothesis that glacial–interglacial cycles are traceable from genetic footprints retained within plant species. Fourteen populations belonging to the Delphinium naviculare species group were sampled from the Tianshan Mountains in western China, and 14 genotypes were detected based on sequences of the two chloroplast fragments trnS–trnG and trnL–trnF. A Bayesian relaxed molecular clock approach revealed that genetic divergence in this species group began in the early Pleistocene and intensified during mid-Pleistocene. A lineage-through-time (LTT) plot shows dramatic accumulations of Delphinium lineages during the three Pleistocene interglacial periods in this area. From ecological niche modeling (ENM), it appears that the distribution of the species group most likely shrunk in the eastern arid and southwestern high-altitude parts of its range during the Last Glacial Maximum (LGM), suggesting that cold and drought were primary drivers of lineage migration during glacial periods. Humid valleys at mid-elevation served as refugia at these times; however, during the latest two glacial epochs, new refugia likely appeared, corresponding to a decreased extent of glaciation. In accordance with the glacial chronology derived from many proxy data such as glacial tills, lacustrine sedimentary sequences, and oxygen isotope records, the genetic structure of the D. naviculare species group reveals a paleoclimatic history that is able to differentiate three glacial–interglacial cycles and an overall trend towards aridity during the course of the Pleistocene.

THE RELEVANCE OF GENE FLOW IN METAPOPULATION DYNAMICS OF AN OCEANIC ISLAND ENDEMIC, OLEA EUROPAEA SUBSP. GUANCHICA

Theoretical and empirical studies suggest that geographical isolation and extinction-recolonization dynamics are two factors causing strong genetic structure in metapopulations, but their consequences in species with high dispersal abilities have not been tested at large scales. Here, we investigated the effect of population age structure and isolation by distance in the patterns of genetic diversity in a wind-pollinated, zoochorous tree (Olea europaea subsp. guanchica) sporadically affected by volcanic events across the Canarian archipelago. Genetic variation was assessed at six nuclear microsatellites (nDNA) and six chloroplast fragments (cpDNA) in nine subpopulations sampled on four oceanic islands. Subpopulations occurring on more recent substrates were more differentiated than those on older substrates, but within-subpopulation genetic diversity was not significantly different between age groups for any type of marker. Isolation-by-distance differentiation was observed for nDNA but not for cpDNA, in agreement with other metapopulation studies. Contrary to the general trend for island systems, between-island differentiation was extremely low, and lower than differentiation between subpopulations on the same island. The pollen-to-seed ratio was close to one, two orders of magnitude lower than the average estimated for other wind-pollinated, animal-dispersed plants. Our results showed that population turnover and geographical isolation increased genetic differentiation relative to an island model at equilibrium, but overall genetic structure was unexpectedly weak for a species distributed among islands. This empirical study shows that extensive gene flow, particularly mediated by seeds, can ameliorate population subdivision resulting from extinction-recolonization dynamics and isolation by distance.

Detection of plant species-specific dna (barley and soybean) in blood, muscle tissue, organs and gastrointestinal contents of rabbit.

The aim of this study was to detect plant DNA sequences from low copy number genes of barley grain and soybean meal, the latter being subjected to solvent extraction process, in blood, liver, kidney, spleen, muscle tissue and digesta (duodenum, caecum and faeces from rectal ampulla) of rabbits. For fattening, Hyla rabbits (20 males and 20 females) were fed a diet including barley grain (15%) and soybean meal (12%). Animals were slaughtered at 74 d of age (2 ± 0.2 kg live weight) and samples collected from each animal. The quality of each DNA sample was verified using the UNIV P/Q primers used to amplify a mammalian specifi c portion of mtDNA 16S rRNA gene. The presence of plant DNA was subsequently ascertained on the same DNA samples, as well as on barley and soybean (control). Two classes of plant DNA sequences were monitored via real-time PCR, using SYBR® Green I Dye: a high copy number chloroplast gene (trnl) and a low copy number specific for barley (metal-dependent hydrolase-like protein) and soybean (lectin) genes. Melting curve analysis was used to identify the PCR products. The chloroplast fragment detection frequency was higher (P<0.01) in muscle (90%), liver (80%), kidney (80%) and spleen (80%) than in blood (40%) and digesta samples. In the latter, chloroplast DNA was found in 40 and 30% of duodenum and caecum contents respectively, and in 30% of faeces. The specificity of the amplicons obtained was checked by sequencing and annotation. In the samples positive for chloroplast fragments, the frequency of detection of barley specific sequence was higher (P<0.01) in liver (62.5%), kidney (62.5%), spleen (62.5%) and digesta (100%) than in blood (25%) and muscle (22.2%) samples. The soybean lectin gene was not detected in animal samples, although it was seen in plant samples. Results confirm that, except for gastrointestinal tract (GIT), plant single copy genes are more difficult to identify in animal samples.

Impacts of recent cultivation on genetic diversity pattern of a medicinal plant, Scutellaria baicalensis (Lamiaceae)

BackgroundCultivation of medicinal plants is not only a means for meeting current and future demands for large volume production of plant-based drug and herbal remedies, but also a means of relieving harvest pressure on wild populations. Scutellaria baicalensis Georgi (Huang-qin or Chinese skullcap) is a very important medicinal plant in China. Over the past several decades, wild resource of this species has suffered rapid declines and large-scale cultivation was initiated to meet the increasing demand for its root. However, the genetic impacts of recent cultivation on S. baicalensis have never been evaluated. In this study, the genetic diversity and genetic structure of 28 wild and 22 cultivated populations were estimated using three polymorphic chloroplast fragments. The objectives of this study are to provide baseline data for preserving genetic resource of S. baicalensis and to evaluate the genetic impacts of recent cultivation on medicinal plants, which may be instructive to future cultivation projects of traditional Chinese medicinal plants.ResultsThirty-two haplotypes of S. baicalensis (HapA-Y and Hap1-7) were identified when three chloroplast spacers were combined. These haplotypes constituted a shallow gene tree without obvious clusters for cultivated populations, suggesting multiple origins of cultivated S. baicalensis. Cultivated populations (hT = 0.832) maintained comparable genetic variation with wild populations (hT = 0.888), indicating a slight genetic bottleneck due to multiple origins of cultivation. However, a substantial amount of rare alleles (10 out of 25 haplotypes within wild populations) lost during the course of S. baicalensis cultivation. The genetic differentiation for cultivated group (GST = 0.220) was significantly lower than that of wild group (GST = 0.701). Isolation by distance analysis showed that the effect of geographical isolation on genetic structure was significant in wild populations (r = 0.4346, P < 0.0010), but not in cultivated populations (r = 0.0599, P = 0.2710). These genetic distribution patterns suggest that a transient cultivation history and the extensive seed change among different geographical areas during the course of S. baicalensis cultivation.ConclusionsAlthough cultivated S. baicalensis maintains comparable genetic diversity relative to wild populations, recent cultivation has still imposed profound impacts on genetic diversity patterns of the cultivated S. baicalensis populations, i.e., the loss of rare alleles and homogenization of cultivated populations. This study suggests that conservation-by-cultivation is an effective means for protecting genetic resources of S. baicalensis, however, the wild resources still need to be protected in situ and the evolutionary consequences of extensive seed exchange mediated by human being should be monitored carefully.

A multi‐locus phylogeny of Euryops (Asteraceae, Senecioneae) augments support for the "Cape to Cairo" hypothesis of floral migrations in Africa

AbstractWith about 100 species, Euryops (Cass.) Cass. ranks among the most speciose genera of the tribe Senecioneae (Asteraceae). The genus has its greatest diversity in South Africa, and displays an interesting disjunct distribution with most of the taxa found in southern Africa and a group of eight endemic species confined to the mountains of tropical East Africa and northeastern Africa. Molecular phylogenetic analyses of DNA sequence data from three chloroplast fragments and the nuclear ITS region were used to reconstruct the evolutionary history of 41 Euryops species in order to unravel species relationships and to determine the origin of the disjunct Afromontane taxa. Our results show a lack of support and resolution in the internal structure of the trees, but also reveal strong incongruence between the ITS and cpDNA datasets as assessed by Bayes Factors. We hypothesise that this is a consequence of the isolation and divergence of many populations over a short time period at some point in the history of the genus. Molecular dating based on our phylogenetic tree suggests that the genus diversified in South Africa around four million years ago. The origin of the East African species, dated at 1.9 Ma, well after the uplift of the East African mountains, is consistent with a scenario of a single dispersal event from South Africa northwards into the tropical East African mountains where diversification occurred, creating a monophyletic group of regional Afromontane endemics.

Fate of transgenic DNA and evaluation of metabolic effects in goats fed genetically modified soybean and in their offsprings

The presence of DNA fragments in blood and milk from goats fed conventional (control) or Roundup Ready® soybean meal solvent extracted (s.e.; treated) was investigated by using a polymerase chain reaction approach. The same investigation was carried out on blood, skeletal muscle and organs from kids of both groups fed only dams’ milk until weaning. Moreover, the possible effects on cell metabolism were evaluated by determination of several specific enzymes in serum, heart, skeletal muscle, liver and kidney. Fragments of the multicopy chloroplast (trnL) gene were found in blood and milk samples from goats of both groups. In kids, the chloroplast fragments were found in samples of both groups. In samples, which proved positive for the presence of chloroplast DNA, fragments of the specific soybean single copy gene (lectin) were detected in several blood and milk samples. The same fragment was also found in control and treated groups of kids. Transgenic fragments were not found in those samples, which were found positive for chloroplast fragments of control groups of either goats or kids. On the contrary, in blood and milk of treated goats, fragments both of the 35S promoter and the CP4 epsps gene were detected. These fragments were also found in treated kids with a significant detection of the 35S promoter in liver, kidney and blood, and of the CP4 epsps gene fragment in liver, kidney, heart and muscle. A significant increase in lactic dehydrogenase, mainly concerning the lactic dehydrogenase-1 isoenzyme was found in heart, skeletal muscle and kidney of treated kids, thus suggesting a change in the local production of the enzyme. Finally, no significant differences were detected concerning kid body and organ weight.

Molecular phylogeny of selected Old World Astragalus (Fabaceae): incongruence among chloroplast trnL‐F, ndhF and nuclear ribosomal DNA ITS sequences

This study reports maximum parsimony and Bayesian phylogenetic analyses of selected Old World Astragalus using two chloroplast fragments including trnL‐F and ndhF and the nuclear ribosomal internal transcribed spacer (nrDNA ITS). A total of 52 taxa including 34 euploid Old World and New World Astragalus, one aneuploid species from the Neo‐Astragalus clade as a representative and 14 other Astragalean taxa, plus Cheseneya astragalina and two species of Caragana as outgroups were analyzed for both trnL‐F and nrDNA ITS regions. ndhF was analyzed in 30 taxa and the same number for the combination of these three datasets were examined. In general, the trnL‐F dataset and the ndhF and nrDNA ITS datasets generated more or less the same clades within Astragalus. However, in the trnL‐F and ndhF phylogenies, Astragalus species are not gathered in a single clade, the so‐called Astragalus s.s., as indicated by the nrDNA ITS tree. Visual inspection of these three phylogenies revealed that they were inconsistent regarding the position and relationships of Astragalus hemsleyi, A. ophiocarpus, A. annularis–A. epiglottis/Astragalus pelecinus,A. echinatus and A. arizonicus. Incongruence length difference test suggested that the trnL‐F, ndhF and nrDNA ITS datasets were incongruent. In spite of this, phylogenetic analyses of the combined datasets as one unit or as three partitions generated trees that were topologically similar as a mix of the cpDNA and the nrDNA ITS trees. However, the combined dataset provided more resolved and statistically supported clades. The recently described A. memoriosus appeared closely related to A. stocksii (both from sect. Caraganella) based on both trnL‐F and nrDNA ITS sequences.

Phylogeny and biogeography of the rice tribe (Oryzeae): Evidence from combined analysis of 20 chloroplast fragments

Based on sequences of 20 chloroplast fragments, we generated a fully resolved phylogeny of Oryzeae and estimated divergence times of its major lineages as well as explored the historical biogeography of the tribe. Our results (1) confirmed the monophyly of Oryzeae and two-subtribe subdivision; (2) indicated that Maltebrunia, Potamophila and Prosphytochloa were genetically distinct enough to deserve generic status but Maltebrunia and Prosphytochloa were sister groups in the subtribe Oryzinae while Potamophila was a member in the subtribe Zizaniinae; (3) suggested that the previously unresolved phylogeny of the subtribe Zizaniinae was most likely explained by insufficient data due to rapid radiation; (4) provided the first well-corroborated timescale for the origin and divergence of Oryzeae, with the crown node of Oryzeae and the deepest split of Oryza at approximately 24 and 15MYA, respectively; (5) developed a biogeographic history of the tribe and substantiated the important roles of long-distance dispersal in the origin and diversification of the tribe Oryzeae.

Phylogeography and host race differentiation in the European mistletoe (Viscum album L.)

Mistletoes are bird dispersed, hemi-parasitic shrubs infecting a large number of woody host plants. Chloroplast fragment length polymorphisms were used to study genetic differentiation among presumed host races, population genetic structure, and to elucidate the postglacial migration history of mistletoe (Viscum album) across the entire natural distribution range in Europe. The populations sampled belong to four closely related taxa, three of which are widely distributed and differ in their host trees, whereas a fourth taxon is rare and endemic to the Island of Crete. The molecular analysis of chloroplast DNA variation supported the distinction of these four taxa. We further found evidence for phylogeographical structure in each of the three widely distributed host races. Independent of host race, mistletoe haplotypes from Turkey were distinct and distant from those found elsewhere in Europe, suggesting that highly differentiated populations, and possibly new taxa, exist at the range limit of the species.

Interlaboratory Transfer of a Real-Time Polymerase Chain Reaction Assay for Quantitative Detection of Genetically Modified Maize Event TC-1507

A real-time polymerase chain reaction (QPCR) assay was developed for quantitative detection of a genetically modified (GM) maize event TC-1507 and modification to conventional PCR for qualitative purposes. Sequences 5'-flanking TC-1507 full-length insert were characterized and showed multiple rearrangements involving insert and maize chloroplast fragments. The event specificity of the TC-1507 assays was based on the detection of transgene and plant rearranged sequences found to 5' flank the insertion site. They were fully specific and exhibited a limit of detection below 10 target copies, allowing consistent detection of 0.1% GM levels. The QPCR was highly linear and efficient and proved adequate for quantification of GM contents, aiming at the fulfillment of legal requirements established in the European Union (i.e., compulsory labeling of TC-1507 levels >0.9%). It satisfactorily determined TC-1507 contents on different matrixes and was successfully transferred a different laboratory.

Phylogenetic relationships and generic delimitation in subtribe Arctotidinae (Asteraceae: Arctotideae) inferred by DNA sequence data from ITS and five chloroplast regions

Asteraceae are the largest family in southern Africa. Elucidating its origins and radiation in the region requires well-supported species-level phylogenies of the lineages. This paper presents a phylogenetic framework for subtribe Arctotidinae, which have a southern and eastern African-Australian distribution centered in the winter-rainfall region of South Africa. DNA sequence data from five chloroplast fragments (ndhF, psbA-trnH, rps16, trnS-trnfM, and trnT-trnF) and the nuclear ITS region were analyzed separately and in combination using parsimony and Bayesian methods. The data sets comprised exemplars from 18 ingroup species, representing the five currently accepted genera, and four outgroup species from Gorteriinae. All analyses indicated Arctotis and Haplocarpha are polyphyletic as presently circumscribed. The Australian-endemic Cymbonotus lawsonianus was placed within a strongly supported clade also containing A. arctotoides from South Africa and H. schimperi from eastern Africa. Retention of Dymondia and resurrection of Landtia at generic level are strongly supported. The phylogenetic hypotheses indicate the subtribe might have originated in temperate southern or eastern Africa, or it was ancestrally widespread in southern Africa and has diversified vicariously. The derived placement of C. lawsonianus indicates long-distance dispersal from southern Africa to Australia occurred.

Genetic analysis of archaeological wood remains: first results and prospects

A total of 51 ancient oak wood samples originating from various European archaeological sites, dating from the Neolithic period to the 18th century, were assayed for the presence of reproducible chloroplast (cp) DNA sequences. Five polymorphic chloroplast fragments were targeted. Only five of the samples could be fully genetically characterised, revealing four different oak cpDNA haplotypes. In all cases, the haplotypes detected on ancient woods and the haplotypes characterised from fresh samples from the same localities matched. Overall, this congruence is consistent with a genetic continuity between ancient and modern European oaks, confirming the hypothesis that the mapped genetic patterns largely reflect the original structure that established during the post-glacial. This stability of the genetic structure implies that, in the future, the technique could be used to infer or confirm the transport of wood by man, providing interesting perspectives for the genetic analysis of ancient woods.