cpDNA trnL Intron Research Articles

Molecular characterization of Dittrichia viscosa (L.) greuter (Asteraceae) populations revealed by ISSR markers and chloroplast (CPDNA) tRNL intron sequences

In this study, molecular characterization of Dittrichia viscosa (L.) Greuter populations collected from Ayd?n province was carried out based on ISSR technique and chloroplast DNA trnL intron sequences. 10 ISSR primers were used to determine the molecular characterization among the populations. For cpDNA trnL intron amplification, trnC and trnD primers were used. In ISSR analysis, a total of 70 bands were obtained. The polymorphism rate was determined to be approximately 94.28%. According to the ISSR analysis, the UPGMA dendrogram consisted of three groups. For cpDNA trnL intron sequences, phylogenetic analyses were obtained along with genetic distances. For populations, cpDNA trnL intron sequences were determined between 454 and 472 bases. The maximum likelihood phylogenetic tree consist of two clades. In addition, cpDNA trnL intron sequences of some species (Chrysophthalmum Pulicaria Inula Jasonia Stenachaenium Carpesium Blumea Iphiona Limbarda Rhanterium Lifago Duhaldea Chiliadenus) of Asteraceae family from NCBI with Dittrichia viscosa populations phylogenetic tree was constructed. As a result of the study, it was determined that the polymorphism rate of Dittrichia viscosa populations obtained by ISSR-PCR was higher than the results obtained from trnL intron sequences.

Sequence Analysis of the Internal Transcribed Spacer (ITS) Region of the Nuclear Ribosomal DNA (nrDNA), and trnL Intron of Chloroplast DNA (cpDNA) of the Chrysochamela (Fenzl) Boiss. (Brassicaceae) In Turkey

Brassicaceae family is an important one since it includes many economic and significant industrial oilseeds, spices, vegetables and some forage plants. In this study, sequences analysis among Chrysochamela (Brassicaceae) species distributed in Turkey were conducted nrDNA ITS and cpDNA trnL intron sequences. Chrysochamela species were collected and brought to the laboratory. ITS and trnL intron sequences were corrected with BioEdit and FinchTV programs. As a result of the study, ITS nucleotide compound compositions were determined as 22.7% T, 29.1 C, 21.5% A and 26.7% G. The lowest distance was 0.000 and the highest distance was 0.037. The phylogenetic tree obtained using the MEGA 6.0 program consists of two large groups. According to trnL intron sequences 37.9% T, 18.4 C, 28.3% A and 15.5% G. Nucleotide compound compositions were determined. The genetic distance between species was determined between 0.000 and 0.022. Maximum likelihood phylogenetic tree consists of two large groups. As a result, phylogenetic analyzes using ITS and trnL intron sequences were compatible with each other. It was also in past studies found to be supported by morphological, anatomical and RAPD data.

Phylogenetic analysis using SCoT markers and chloroplast trnL intron in some Eriobotrya japonica (Thunb.) Lindl. (Rosaceae) populations from the Aegean region of Turkey

In this study, phylogenetic analyses of some Eriobotrya japonica (Thunb.) Lindl. populations based on SCoT marker technique and cpDNA trnL intron sequence analysis were performed. Specimens from the populations were collected from the Aegean region of Turkey and brought to the laboratory for genomic DNA isolation. To determine the phylogenetic analysis of Eriobotrya japonica populations, seven SCoT primers were used. In the SCoT analysis, a total of 54 bands were obtained, 49 of which were polymorphic and 5 were monomorphic. In chloroplast trnL intron region, trnC, trnD primers were used for PCR amplification. Eriobotrya japonica trnL intron sequence lengths were determined to be between 530 and 547 nucleotides. The average nucleotide rate detected as 38.9% for thymine, 18.1% for cytosine, 28.2% for adenine and 14.8% for guanine. Rosaceae family trnL intron sequences belonging to Rhaphiolepis Lindl., Heteromeles M.Roem., Photinia Lindl. Cotoneaster Medik, Sorbus L., Malus Miller, Pyrus L., Prunus L. and Rosa L. were retrieved from NCBI and their phylogenetic relationship with Eriobotrya japonica populations were revealed. As a result of the study, polymorphism rate was determined as 90.74% in SCoT analysis. According to cpDNA trnL intron results, Eriobotrya japonica populations were identified in the same group with Rhaphiolepis indica, a finding supported by past phylogenetic analyzes.

Molecular identification and analysis of Psidium guajava L. from indigenous tribes of Taiwan

Psidium guajava L. is a perennial fruit tree in subtropical and tropical areas. In Taiwan, P. guajava has been used as anthro-pharmacological plants by aboriginal people to treat acute diarrhea, cough and intestinal spasmodic diseases. The classification and functional identification of P. guajava remains unsolved these days. In this study, molecular markers 18S rRNA, internal transcribe spacer (ITS) region of ribosomal DNA, trnL intron and trnL-trnF intergenic spacer (IGS) of chloroplast DNA (cpDNA), and random amplified polymorphic DNA (RAPD) were used for the molecular identification of 18 P. guajava samples from different indigenous tribes, 2 from non-indigenous tribe, and 12 commercial cultivars from markets in Taiwan. Molecular methods restricted fragment length polymorphism (RFLP) and denatured gradient gel electrophoresis (DGGE) are found time-consuming and less efficient as compared to RAPD, thus are not suitable for samples of high homology. In this study, 18S rDNA, ITS and cpDNA trnL intron and trnL-trnK intergenic spacer were also tested molecular marker; however, results analyzed by molecular algorithm UPGMA, Neighbor-Joining, Parsimony or Maximum likelihood showed no discriminations (data not shown). On the other side, ten 10-mer oligonucleotide primers were used in RAPD to amplify the specific genes from 32 guava samples. Four primers, OPB 17, OPG 6, OPY 15 and OPY 18, were able to direct the amplification and yielded a total of 82 polymorphic RAPD patterns. Thirty-two genotypes on the dendrogram were identified and were divided into two major groups, the uncultivated and commercial cultivars. Based on the cluster analysis, the red-flesh Psidium samples that were believed to have high medical function were grouped independently. The results suggest that RAPD is useful for the discrimination of uncultivated, cultivars and potential Psidium of high economy.

Genetic Diversity and Phylogenetic Analyses of Laurus nobilis L. (Lauraceae) Populations Revealed Chloroplast (cpDNA) trnL Intron and trnL-F Region

ABSTRACT In this study, genetic diversity and phylogenetic analyzis based on cpDNA trnL intron and trnL-F sequences of L. nobilis L. populations were made. Persea schiedeana and Endiandra wolfei taxa were used as outgroups. Plant samples were collected from various different provinces and districts of the Aegean region and brought to the laboratory. Genomic DNAs were isolated from fresh and green leaves of the plants. trnC and trnD primers were used for the trnL intron region, and trnE and trnF primers were used for the trnL-F region. The obtained DNA sequences were then corrected using BioEdit and FinchTV programs. MEGA 6.0 program was used both phylogenetic analyzis and the detection of genetic distances. For the intron region of the trnL, the mean nucleotide composition was determined as 36.5% T, 20.2% C, 26.4% A and 16.9% G. These values were found as 30.7% T for 15.9% C, 31.9% A and 21.6% G for the trnL-F region. The lowest distance for the trnL intron region was found 0.000 while the highest distance was 0.004. The value was found 0.000 for the trnL-F region. The phylogenetic tree, which was obtained using both trnL intron and trnL-F sequences, consisted of two large branches. As a result, according to the data obtained from trnL intron and trnL-F sequences, the trnL intron sequence found a solvent in the phylogenetic relationship between L. nobilis populations, while the phylogenetic relationship between trnL-F sequence and L. nobilis populations was not found.

Molecular phylogenetic studies based on rDNA ITS, cpDNA trnL intron sequence and cladode characteristics in nine Protasparagus taxa.

The genus Asparagus comprises three subgenera of cladode bearing plants: Protasparagus, Asparagus, and Myrsiphyllum. The interspecific delimitation of the subgenus Protasparagus is ill-defined till date. In the present study, interspecific phylogenetic relationships among nine taxa of Protasparagus based on ribosomal DNA internal transcribed spacer region (ITS1-5.8S-ITS2) sequence and the chloroplast DNA trnL intron sequence conservation with their cladode morphology, anatomy, and stomatal characteristics have been analyzed for the first time. The monophyletic subgenus Protasparagus could be resolved into four strongly supported distinct subclades (I, II, III and IV) suggesting that the rDNA and cpDNA molecular phylogenies are explicitly congruent with the cladode characteristics of the subgenus Protasparagus. The present study also confirms the existing subgeneric classification of the genus Asparagus with the monophyletic origin of the dioecious subgenus Asparagus. The present work brings out phylogenetic and taxonomic relationships within the studied taxa of the subgenus Protasparagus therefore providing important background information for further studies on biogeography of a wide range of species.

Comparison of methods for rooting phylogenetic trees: A case study using Orcuttieae (Poaceae: Chloridoideae)

DNA sequence data (cpDNA trnL intron and nrDNA ITS1 and ITS2) were analyzed to identify relationships within Orcuttieae, a small tribe of endangered grasses endemic to vernal pools in California and Baja California. The tribe includes three genera: Orcuttia, Tuctoria, and Neostapfia. All three genera carry out C 4 photosynthesis but aquatic taxa of Orcuttia lack Kranz anatomy. The unusual habitat preference of the tribe is coupled with the atypical development of C 4 photosynthesis without Kranz anatomy. Furthermore, the tribe has no known close relatives and has been noted to be phylogenetically isolated within the subfamily Chloridoideae. In this study we examine the problem of inferring the root of the tribe in the absence of an identified outgroup, analyze the phylogenetic relationships of the constituent taxa, and evaluate the evolutionary development of C 4 photosynthesis. We compare four methods for inferring the root of the tree: (1) the outgroup method, (2) midpoint rooting, the imposition of a molecular clock for both (3) maximum likelihood (ML) and (4) Bayesian analysis. We examine the consequences of each method for the inferred phylogenetic relationships. Three of the methods (outgroup rooting and the ML and Bayesian molecular clock analyses) suggest that the root of Orcuttieae is between Neostapfia and the Tuctoria/ Orcuttia lineage, while midpoint rooting gives a different root. The Bayesian method additionally provides information about probabilities associated with other possible root locations. Assuming that the true root of Orcuttieae is between Neostapfia and the Tuctoria/ Orcuttia lineage, our data indicate Neostapfia and Orcuttia are both monophyletic, while Tuctoria is paraphyletic (with no synapomorphies in either dataset) and forming a grade between the other two genera and needs taxonomic revision. Our data support the hypothesis that Orcuttieae was derived from a terrestrial ancestor and evolved specializations to an aquatic environment, including C 4 photosynthesis without Kranz anatomy.

Molecular phylogeny and biogeography of the bipolar Euphrasia (Orobanchaceae): Recent radiations in an old genus

Euphrasia includes perennial or annual green parasitic plants, and has a disjunct bipolar distribution except for one transtropical connection across the high mountains of Oceania. The disjunction is coupled with strikingly contrasting patterns of morphological diversity between the southern and northern hemispheres, making it an exciting model to study processes of evolutionary diversification which shaped present floras. We inferred the relationships among 51 species representing 14 of the 15 sections of the genus based on nrDNA ITS and cpDNA trnL intron, trnL- trnF and atpB- rbcL intergenic spacers. Maximum parsimony and Bayesian inference support monophyly of the genus and of several intrageneric groups characterized by morphology, ploidy level, and geographic range. Molecular phylogenetic dating using Bayesian “relaxed” clock methods suggests that the earliest Euphrasia radiations occurred minimum 11–8 Mya with bipolarity being achieved 7–5 Mya. Biogeographic analyses using explicit model-based approach inferred Eurasia as an ancestral area for the genus. The most parsimonious reconstruction found by a dispersal-vicariance analysis requires 17 dispersals to account for the current biogeographic pattern and supports Eurasian origin for Euphrasia. Both long-distance dispersal and across land vicariance can be invoked to explain the diversification in the genus, which experienced rapid radiations driven by new ecological opportunities of the late Pliocene and Pleistocene but also retained a set of local endemic or relict species of an earlier origin.

Nomenclature in the dock. Overclassification leads to instability: a case study in the horticulturally important genus Cyclamen (Myrsinaceae)

Nine different classifications have been produced in the last 70 years for the horticulturally valuable genus Cyclamen, a small genus with fewer than 30 species. These classifications, generated by intuitive methods and cladistic analyses, incorporated a total of four infrageneric ranks above that of species and were based on data from morphology, cytology and DNA sequencing. Our results, based on cladistic analyses of three independent data sources − nrDNA ITS, cpDNA trnL intron and morphological data − reveal good resolution only in nrDNA sequence data. However, when these three data sources are combined they provide stronger resolution and support for three major clades, only one of which, subgenus Psilanthum, has been consistently supported in previous classifications. The differing infrageneric classifications produced in Cyclamen result from varying taxon sampling, differing interpretation of morphological data, changes in the sources and analysis of data, and inconsistent application of names. Extensive subdivision of small genera in the absence of adequate data that could provide evidence for consistent patterns of relationship is premature and leads to a proliferation of names.© 2004 The Linnean Society of London, Botanical Journal of the Linnean Society, 2004, 146, 339-349.

The tribal position of Fockea and Cibirhiza (Apocynaceae: Asclepiadoideae): evidence from pollinium structure and cpDNA sequence data

The pollinium morphology of the two members of the Asclepiadoideae, tribe Fockeeae, Fockea Endl. and Cibirhiza Bruyns, has been studied in detail and compared with that of eight genera of Marsdenieae, the tribe in which Fockea and Cibirhiza were previously accommodated and thus their putative closest relatives, as well as nine genera of Asclepiadeae. Both Fockea and Cibirhiza have several morphological characteristics in common, the most important of which is the absence of well-developed caudicula, which distinguishes them from all other genera of Asclepiadoideae known. The pollinium structure of these two genera, however, differs significantly. Whereas the pollinium of Cibirhiza consists of single pollen grains and is covered by a pollinium wall, as is typical for other Asclepiadoideae, the pollinium of Fockea consists of tetrads and is not covered by a pollinium wall, a condition otherwise typical of Secamonoideae. Fockea, however, has only two pollinia per anther, as does Cibirhiza and all other Asclepiadoideae, whereas the Secamonoideae have four pollinia per anther. Sequence data from two intergenic spacers, trnT-L and trnL-F and the trnL intron of cpDNA was analyzed. The ingroup included three species of Fockea and one species of Cibirhiza. The outgroup taxa consisted of three representatives each of Periplocoideae, and Secamonoideae and 24 species of Asclepiadoideae, including representatives of all tribes, of which eight genera belong to Marsdenieae, as outgroups. The results of the DNA analysis provide strong support for Fockeeae as a monophyletic tribe, distinct from Marsdenieae and, to the rest of the Asclepiadoideae. With the exception of pollen data, all morphological and molecular evidence clearly support recognition of the tribe Fockeeae. The occurrence of two such significantly different types of pollinia structure – characters elsewhere in the family used to distinguish subfamilies – within the small tribe Fockeeae was unexpected, and can perhaps best be understood as yet another attestment to the basal position of the Fockeeae in the nascence of the Asclepiadoideae.

A molecular phylogeny and generic rearrangement of the stapelioid Ceropegieae (Apocynaceae-Asclepiadoideae)

Representatives of nearly all genera of the taxon-rich stem-succulent stapeliads and most of the few related, leafy genera were analyzed. Sequence data from two non-coding molecular markers (ITS region of nrDNA and trnT-L and trnL-F spacers as well as the trnL intron of cpDNA) support the traditional tribal affiliation of the genera, which form a monophyletic group. This monophylum breaks into a basal Neoschumannia/Anisotoma/Riocreuxia/Sisyranthus nk;clade, from which the core Ceropegieae are derived. The four Ceropegia species included are not monophyletic, and their relationship to Brachystelma changes depending on the marker studied. The stem succulent taxa fall in a number of well supported, but unresolved clades, the most prominent being the predominantly southern African clade comprising Orbea, Stapelia and some other genera. The most derived taxa of NE Africa, Duvaliandra and White-sloanea, are basal to this southern African clade. The other clades comprise the more basal genera of stem-succulent stapeliads, including the members of the Caralluma complex. Of the 17 genera accepted by Plowes for the Caralluma complex, seven are recognized: Caralluma, Apteranthes, Australluma, Boucerosia, Caudanthera, Desmidorchis and Monolluma. New combinations are proposed in 15 cases; Caralluma adscendens var. geniculata is raised to specific rank. Anomalluma is reinstated, and Pseudolithos mccoyi is transfered to it. A broadened concept for Orbea (incl. Angolluma and Orbeopsis) is recognized, but Orbeanthus is kept separate. The monotypic Ballyanthus, recently separated from Orbea, is nested within Duvalia. Piaranthus (incl. Huerniopsis) is monophyletic. The bitypic Notechidnopsis is reduced to the type species, N. tessellata, while N. columnaris is transferred to a new genus, Richtersveldia.

Chloroplast DNA phylogeny and biogeography of Lepidium (Brassicaceae)

Two intergenic spacers, trnT-trnL and trnL-trnF, and the trnL intron of cpDNA were sequenced to study phylogenetic relationships and biogeography of 73 Lepidium taxa. Insertions/deletions of ≥3 bp (base pairs) provided reliable phylogenetic information whereas indels ≤2 bp, probably originating from slipped-strand mispairing, are prone to parallelism in the context of our phylogenetic framework. For the first time, an hypothesis of the genus Lepidium is proposed based on molecular phylogeny, in contrast to previous classification schemes into sections and greges (the latter category represents groups of related species within a given geographic region), which are based mainly on fruit characters. Only a few of the taxa as delimited in the traditional systems represent monophyletic lineages. The proposed phylogeny would suggest three main lineages, corresponding to (1) sections Lepia and Cardaria, (2) grex Monoplocoidea from Australia, and (3) remaining taxa, representing the bulk of Lepidium species with more or less resolved sublineages that sometimes represent geographical correspondence. The fossil data, easily dispersible mucilaginous seeds, widespread autogamous breeding systems, and low levels of sequence divergence between species from different continents or islands suggest a rapid radiation of Lepidium by long-distance dispersal in the Pliocene/Pleistocene. As a consequence of climatic changes in this geological epoch, arid/semiarid areas were established, providing favorable conditions for the radiation of Lepidium by which the genus attained its worldwide distribution.

Inclusion of Tenaris and Macropetalum in Brachystelma (Apocynaceae-Asclepiadoideae-Ceropegieae) inferred from non-coding nuclear and chloroplast DNA sequences

The inclusion of Tenaris and Macropetalum in Brachystelma as proposed by Peckover in 1996 and contested by Victor and Nicholas in 1998 is supported by molecular studies. Parsimony analysis of sequence data from two non-coding molecular markers (ITS region of nrDNA and trnT-L and trnL-F spacers as well as the trnL intron of cpDNA) suggests a well-supported Brachystelma s.l. clade (including Tenaris and Macropetalum) with little internal resolution. The Brachystelma s.l. clade occupies a sistergroup position to the Ceropegia/stapeliad clade, and both clades together are sister to an Anisotoma/Sisyranthus/Neoschumannia clade.

RELATIONSHIPS AND DIVERGENCE PATTERNS IN HYPOPTERYGIUM ‘ROTULATUM’ S.L. (HYPOPTERYGIACEAE, BRYOPSIDA) INFERRED FROM trnL INTRON SEQUENCES. STUDIES IN AUSTRAL TEMPERATE RAIN FOREST BRYOPHYTES 7

The phylogenetic relationship of species within the Hypopterygium ‘rotulatum’ (Hedw.) Brid. s.l. complex is analysed by sequencing the trnL intron of cp DNA. Furthermore, the divergence in variable New Zealand H. ‘rotulatum’ samples is investigated along a latitudinal gradient. The presented data strongly support the delimitation of at least two distinct species within Hypopterygium ‘rotulatum’ s.l. in New Zealand, Australia and Tasmania, which correspond with the morphological species H. muelleri Hampe (possibly conspecific with H. debile Reichardt) and H. didictyon Müll. Hal., a species known from Chile. The distribution patterns of these species are discussed on a regional (New Zealand) and global scale; indicating an australasian distribution pattern of H. muelleri and a palaeoaustral pattern of H. didictyon.

Phylogenetic Use of Noncoding Regions in the GenusGentianaL.: ChloroplasttrnL (UAA) Intron versus Nuclear Ribosomal Internal Transcribed Spacer Sequences

Sequence divergence was estimated within noncoding sequences of both chloroplast DNA (cpDNA)trnL (UAA) intron and nuclear ribosomal DNA (nrDNA) internal transcribed spacer sequences (ITS1 and ITS2) for 10 species of the genusGentianaL. (Gentianaceae). Comparisons of evolutionary rates among these sequences (cpDNA versus nrDNA, ITS1 versus ITS2) were performed. It appears that sequence divergence is on average two to three times higher in ITSs than in thetrnL intron sequences and higher in ITS1 than in ITS2. Both the cpDNA intron and ITSs of nrDNA give concordant phylogenetic trees. However, the ITS-based phylogeny displays higher bootstrap values. At the intrageneric level, at least inGentiana,ITSs (especially ITS2) sequences seem to be more appropriate in the assessment of plant phylogenies. Nevertheless, the cpDNAtrnL intron seems to be preferable at the intergeneric level.