psbA-trnH Intergenic Spacer Research Articles

Variability of Chloroplast DNA in Oxytropis Section Polyadena (Fabaceae) from Asian Russia: Population Analysis and Phylogenetic Relationships

Nucleotide polymorphism of the intergenic spacers psbA–trnH, trnL–trnF, and trnS–trnG of chloroplast DNA was analyzed in populations of six Oxytropis species of the section Polyadena from Asian Russia. A low level of nucleotide diversity (0.0001–0.0014) was found, except for two O. glandulosa populations (0.0036 and 0.0059). It was noted that the haplotype diversity varies from 0.133 to 0.911. It was established that high interspecific genetic distances and detected species-specific nucleotide substitutions and indels indicate a significant differentiation of the chloroplast genomes of O. muricata, O. microphylla, O. trichophysa, and O. glandulosa (lineages 2, 3). The species O. pseudoglandulosa, O. muricata, O. varlakovii, and O. glandulosa (lineage 1) form a weakly differentiated complex, which is probably caused by their relatively recent divergence.

An integrative approach reveals five new species of highland papayas (Caricaceae, Vasconcellea) from northern Peru.

The assignment of accurate species names is crucial, especially for those with confirmed agronomic potential such as highland papayas. The use of additional methodologies and data sets is recommended to establish well-supported boundaries among species of Vasconcellea. Accordingly, six chloroplast (trnL-trnF, rpl20-rps12, psbA-trnH intergenic spacers, matK and rbcL genes) and nuclear (ITS) markers were used to delimit species in the genus Vasconcellea using phylogeny and four DNA-based methods. Our results demonstrated congruence among different methodologies applied in this integrative study (i.e., morphology, multilocus phylogeny, genetic distance, coalescence methods). Genetic distance (ABGD, SPN), a coalescence method (BPP), and the multilocus phylogeny supported 22-25 different species of Vasconcellea, including the following five new species from northern Peru: V. badilloi sp. nov., V. carvalhoae sp. nov., V. chachapoyensis sp. nov., V. pentalobis sp. nov., and V. peruviensis sp. nov. Genetic markers that gave better resolution for distinguishing species were ITS and trnL-trnF. Phylogenetic diversity and DNA-species delimitation methods could be used to discover taxa within traditionally defined species.

<p><strong><em>Allium gilanense</em>, a new species of <em>Allium</em> sect. <em>Codonoprasum</em> (Amaryllidaceae) from Iran: evidence from morphological and molecular data</strong></p>

As a result of recent botanical expeditions to the north of Iran, we describe here a new endemic Allium species from Gilan province named Allium gilanense. Molecular and morphological data indicate that it belongs to Allium sect. Codonoprasum. We provide a morphological description, comparing Allium gilanense with the closest relative taxa A. lenkoranicum and A. paniculatum, the preliminary karyotype of the new species, and molecular phylogenetic data derived from the nuclear ribosomal DNA internal transcribed spacers (ITS) and the chloroplast intergenic spacer trnH-psbA. The chromosome number of the new species is 2n = 16 with 0-3 B chromosomes.

Identification of morphologically similar species of Tribulus (Zygophyllaceae) by employing DNA barcodes and rRNA secondary structures

Abstract Tribulus terrestris is one of the essential constituents in traditional Indian systems of medicine and commonly used in several herbal preparations. It is adulterated with morphologically similar species, viz., T. subramanyamii, T. lanuginosus and T. alatus. T. terrestris is often misidentified for T. subramanyamii and T. lanuginosus. Hence, the error-free recognition and collection of this medicinal herb are important to ensure the drug’s effectiveness and biosafety. In the present study, DNA barcoding technique was employed to identify and to differentiate T. terrestris from its closely related species. Phylogenetic analysis was carried out for the three species of Tribulus using barcode candidates, such as, nuclear ribosomal DNA region ITS, chloroplast plastid gene psbA-trnH and rbcL. Sequence alignment revealed 7.5% polymorphic sites in ITS, with different specific regions showing the distribution of polymorphic sites as follows: ITS1 1.7%, ITS2 3.6% and the plastid intergenic spacer psbA-trnH 2.33%. The region ITS2 possesses a higher rate of transition/transversion ratio, percentage of variation and mean distance as compared to ITS1 and psbA-trnH. In addition, ITS2 has unique polymorphic loci which aid to differentiate T. terrestris from other species of Tribulus. Additionally we predicted the structure of ITS2 and 5.8S rRNA which distinguishes three phenotypically alike species based on the helical structure. The present study opens up new avenue for the identification of closely related species based on ITS2 region and 5.8S rRNA structure which could be employed for diversity studies

Generation of novelties in the genus Ocimum as a result of natural hybridization: A morphological, genetical and chemical appraisal

The genus Ocimum is a boutique of a diverse set of specialized metabolites such as terpenoids and phenylpropanoids. Each Ocimum species and its cultivars represent a characteristic chemical profile. The present study explored the two interspecific Ocimum hybrids originating through a serendipitous natural cross between O. kilimandscharicum and O. basilicum. These two novel Ocimum hybrids exhibited intermediate morphological features of two parental species. Inter simple sequence repeats (ISSR) analysis and DNA barcoding with the plastid non-coding trnH-psbA intergenic spacer region reaffirmed unambiguous parental identification and differentiation of these natural hybrids from other available Ocimum species. Consequently, gas chromatography-mass spectrometry-based metabolite profiling of two hybrids identified them as specific chemotypes with the presence of a unique blend of specialized metabolites from the parental species, which are either rich in terpenes or phenylpropanoids. Additionally, expression analysis of key genes from terpenoid and phenylpropanoid pathways corroborated with differential metabolite accumulation. Thus, these two Ocimum hybrids represented the novel chemotypes, which could be useful in commercial cultivation to produce novel essential oil and bioactive constituents. Further, the resulting metabolite diversity could have potential industrial applications in the areas of healthcare, bioremediation, and crop protection.

DNA barcoding supports existence of morphospecies complex in endemic bamboo genus Ochlandra Thwaites of the Western Ghats, India

Ochlandra Thwaites, an economically exploited bamboo genus of the Western Ghats of India is severely affected by unsustainable extraction, natural habitat destruction and endangerment of species resources. This taxonomically challenging genus consists of a genetic mixture of 10 related polyploid species that are difficult to define and classify using traditional morphology. The present study investigated the probability of DNA barcoding using seven standard barcode regions recommended by CBOL as a supplementary tool to define true species boundaries. Distance (MEGA v.6.0) and sequence similarity (TaxonDNA) based approaches highlighted the discriminatory power of psbA-trnH intergenic spacer barcode region, but did not support true species entities. Neighbour-joining and Bayesian inference trees supported the existence of morphospecies complex in seven species of the genus owing to weak reproductive barriers amongnaturally coexisting species. Morphological affinities existing within genus might have stemmed from natural interspecific hybridization events and consequent reticulate evolution in morphospecies complex of genus Ochlandra.

Molecular authentication of the Chinese herb huajuhong and related medicinal material by DNA sequencing and ISSR markers

DNA sequences in the chloroplast (trnH-psbA intergenic spacer) and nuclear (ITS) regions were amplified and determined for Huajuhong derived from the peels of the immature fruits of Citrus grandis 'Tomentosa' and related medicinal material. These sequences together with the inter-simple sequence repeats (ISSR) markers may be used to differentiate C. grandis 'Tomentosa' from other Citrus variants for the prevention of misuse.

Barcoding of ToLCV Resistant Tomato Germplasm in Bangladesh

The current study was carried out to confirm the existence of the ToLCV resistant genes (Ty-1 to Ty-5) in the germplasm using molecular markers and to identify at the genomic level following phylogenetic relationships analysis among some local tomato germplasm using DNA barcoding. Most of the tomato germplasm (Ten out of 14) contain the dominant Ty-genes as revealed by PCR analysis. “Barcoding” of the non-coding plastid trnH-psbA intergenic spacer region, three plastidal regions: rbcL, rpoB, rpoC1, spacer region of nuclear genome ITS and a mitochondrial region matK were employed following PCR and sequence analysis of the germplasm. Among all the barcode genes, rpoB, rbcL, trnH-psbA and ITS were leading candidates for successful amplification and used for the identification of the germplasm as S. lycopersicum in multi-locus identification based on their sequences. Neighbor-Joining phylogenetic tree was constructed in which the germplasm were clustered into five main clades. The current study was successful to establish an efficient barcoding protocol for the correct identification of tomatoes and was capable of establishing elite gene source(s) for biotic stress resistance tomato varieties which would serve as potential donor plants in modern breeding programs.
 Plant Tissue Cult. & Biotech. 30(1): 107-117, 2020 (June)

A multilocus phylogeny of the non-photosynthetic parasitic plant Cistanche (Orobanchaceae) refutes current taxonomy and identifies four major morphologically distinct clades

Phylogenetic relationships of and within non-photosynthetic parasitic lineages are notoriously poorly known, which negatively affects our understanding of parasitic plants. This is also the case for Cistanche (Orobanchaceae), an Old World genus with about two dozen species, whose relationships have not yet been addressed using molecular phylogenetic approaches. Here we infer phylogenetic relationships within the genus, employing a taxonomically and geographically broad sampling covering all previously distinguished infrageneric groups and most of the currently recognized species. A combined matrix of three plastid markers (trnL-trnF, including the trnL intron and the intergenic spacer (IGS), trnS-trnfM IGS and psbA-trnH IGS) and one nuclear marker (ITS) was analyzed using maximum parsimony, maximum likelihood and Bayesian inference. Cistanche falls into four well-supported and geographically differentiated clades: East Asian Clade, Northwest African Clade, Southwest Asian Clade and Widespread Clade. Of those, only the East Asian Clade corresponds to a previously recognized taxonomic section, whereas the others either contain members of two or three sections (Widespread Clade and Southwest Asian Clade, respectively) or have not been taxonomically recognized so far (Northwest African Clade). Whereas the Southwest Asian Clade exhibits strong phylogenetic structure among and partly within species (the East Asian Clade and the Northwest African Clade are monospecific), phylogenetic resolution within the Widespread Clade is often low and hampered by discrepancies between nuclear and plastid markers. Both molecular and morphological data indicate that species diversity in Cistanche is currently underestimated.

Molecular and elemental characterizations and taxonomic notes of three endemic Lathyrus L. species

In this study, the morphological, genetical (molecular) and elemental compositions of morphologically similar three endemic Lathyrus L. species such as L. phaselitanus Hub.-Mor. & Davis, L. lycicus Boiss. and L. belinensis N. Maxted & D. J. Goyder belong to genus Lathyrus in Turkey were investigated. In the morphological part, some important parameters such as stem shape, plant height, leaflet number, leaflet length, leaflet width, stipule shape, stipule length, stipule width, fruit length, fruit width, fruit shape, number of flowers and standard were investigated. In the genetical part of the study, chloroplast intergenic spacer trnH-psbA was used to reveal the genetic diversity and phylogenetic relationship among the three endemic Lathyrus species to decide whether they were systematically similar or not. Additionally, the concentration of some elements such as Nickel (Ni), Iron (Fe), Copper(Cu), Cadmium (Cd), Chromium (Cr), Manganese (Mn), Magnesium (Mg), Zinc (Zn), Sodium (Na) and Potassium (K) were also determined by flame atomic absorption spectrometry to observe the similarities and/or differences among these three endemic Lathyrus species based on elemental composition. The results revealed that there was also a genetic similarity of 99% between morphologically similar L. belinensis and L. lycicus species but they were previously placed in different sections.

<p><strong>Is <em>Rosa × archipelagica</em> (Rosaceae, Rosoideae) really a spontaneous intersectional hybrid between <em>R. rugosa </em>and <em>R. maximowicziana</em>? Molecular data confirmation and evidence of paternal leakage</strong></p>

Rosa × archipelagica is a spontaneous hybrid between Rosa rugosa and R. maximowicziana distributed, together with its parental taxa, on the coast of a small Stenina Island in the Peter the Great Gulf, the Sea of Japan (Russian Far East). Its hybrid origin is confirmed by direct electropherogram comparisons of PCR products and maximum likelihood analyses of nuclear ITS 1–2 and plastid intergenic spacers trnH–psbA and ndhC–trnV. The plastome markers indicate R. maximowicziana as the maternal genome donor of Rosa × archipelagica, however, one specimen shows also signs of heteroplasmy. This is the first evidence of possible biparental inheritance of plastids in rose hybrids. The results do not show whether the hybrid plants all belong to F1 or further generations of hybrids.

A trnH-psbA barcode genotyping assay for the detection of common apricot (Prunus armeniaca L.) adulteration in almond (Prunus dulcis Mill.)

ABSTRACT Almond kernels can be mixed with apricot (Prunus armeniaca L.) seeds while preparing certain almond-based products such as marzipan, without an identifiable alteration in the final product’s sensory attributes. Consequently, apricot kernels are the most commonly utilized almond substitutes for the economic adulteration of almonds. In the present work, the trnH-psbA intergenic spacer was evaluated as a potential PCR-based authenticity marker of almond. The barcode enabled the straightforward discrimination of almond and apricot in their blends, based on diagnostic length polymorphisms between the two closely related Prunus species. The proposed assay simultaneously amplifies and distinguishes almond- and apricot-specific trnH-psbA barcode fragments from mixed samples. The barcode size of almond segregates significantly from peach as well, which is a secondary source for the economic adulteration of almond kernels. The proposed trnH-psbA genotyping assay is easy and selective for the discrimination of almond from its potential adulterants when present as admixtures.

Incongruence of morphological determinations and DNA barcode sequences: a case study in Cinnamomum (Lauraceae)

In the course of molecular systematic studies of Lauraceae we received a sample of a plant cultivated under the name Cinnamomum porrectum in the Botanical Garden Munchen-Nymphenburg. Preliminary determinations, both morphologically based on the Flora of China (Li & al. 2008) and by chloroplast sequences (psbA-trnH spacer, trnK intron including matK gene, trnL intron, trnL-trnF spacer and trnQ-rps16 spacer) obtained by Sanger sequencing suggested that it was C. camphora, still the plant looked different from other individuals of C. camphora cultivated in the botanical gardens of Berlin, Hamburg, Mainz, Munich and Oldenburg. Attempts to sequence the more informative nuclear internal transcribed spacer repeatedly led to mixed signals. We therefore used Illumina sequencing on a set of pre-amplified molecular markers (ITS, trnK 3′ and 5′ intron, trnL intron, and the intergenic spacers psbA-trnH, trnL-trnF, as well as parts of the trnQ-rps16 spacer), and downloaded available sequences of C. camphora and C. parthenoxylon from GenBank for comparison. Considerable differences were found among these sequences, but the haplotype groups do not coincide with the current species determinations. Particularly the internal transcribed spacer sequences are rather diverse, suggesting possible misidentifications, contaminations, and/or a common gene pool that is larger than anticipated. Concerning the plant in question, our results suggest that it may be a hybrid, with C. camphora as the maternal and another species, possibly C. parthenoxylon, as the paternal parent.Citation: Rohwer J. G., Trofimov D., Mayland-Quellhorst E. & Albach D. 2019: Incongruence of morphological determinations and DNA barcode sequences: a case study in Cinnamomum (Lauraceae). – Willdenowia 49: 383–400. doi: https://doi.org/10.3372/wi.49.49309Version of record first published online on 4 December 2019 ahead of inclusion in December 2019 issue.

Barcoding assessment of the Citrus species cultivated in the eastern Afghanistan

The establishment of a modern fruit culture in developing countries requests an accurate evaluation of the preexisting germplasm and its health status. This to prevent the possibility to introduce new germplasm which can be easily prey of the endemic diseases carried by asymptomathic host plants. Therefore, after the identification of cases of citrus plants affected by Tristeza virus, a survey of the germplasm cultivated in the Nangarhar valley and some nearby regions was run. The survey was focussed on the identification of the main Citrus species widely cultivated using barcoding analysis of conserved sequences located in the plastid DNA. The sequences of matK and rbcl genes did not show any discriminatory ability while the analysis of the the non-coding psbA-trnH intergenic spacer (psbA-trnH) showed a robust single nucleotide polymorphism (sNP) discriminating C. aurantium from C. sinensis in all analysed samples. These non-coding regions have no known function; thus, much of the variation may result from the spread of mutations unconstrained by selection. Because nucleotide variation in the psbA-trnH spacer region is high, mutational hot spots may be useful to detect species-level variations. According to our study, the afghan citrus germplasm belong to the C. aurantium species which is commonly used in citrus culture as a rootstock. In Afghanistan it is widely cultivated for fresh consumption, without topworking with selected varieties and this may be the reason why symptoms are often mild and cultivation can be anyhow carried on.

The use of plant DNA barcoding coupled with HRM analysis to differentiate edible vegetables from poisonous plants for food safety

Poisonous Urobotrya siamensis Hiepko has caused deaths of consumers due to misidentification in several cases in past years. The poisonous plant shares the vernacular name in Thai “Phak Wan” with two vegetables, Melientha suavis Pierre and Sauropus androgynus (L.) Merr. The morphology of young leaves among these species is also similar. Therefore, consumer safety could be a major concern. We applied DNA barcoding coupled with high-resolution melting (Bar-HRM) analysis to differentiate the poisonous species from the edible species. Four regions of DNA barcodes, rbcL, matK, ITS2 and the psbA-trnH intergenic spacer, were analysed. Bar-HRM primer pairs were designed based on variable sites of the DNA sequence of the chloroplast matK gene. Distinct melting temperatures for U. siamensis, M. suavis and S. androgynus were revealed at 77.94 ± 0.06 °C, 77.47 ± 0.06 °C and 76.80 ± 0.00 °C, respectively, with confidence levels >99%. The method sensitively identified the poisonous U. siamensis species at levels as low as 0.01 ng of DNA template. Bar-HRM analysis was capable of identifying seedlings or young plants from local nurseries and proved to be an efficient method for detection of food processed by heat treatment, including 5 min of stir-frying and 15 min of boiling. This study is the first report showing that Bar-HRM analysis for safety concerns effectively differentiated poisonous plants from edible plants. Moreover, Bar-HRM analysis may be further applied to other vegetables for quality control, from crop production to food control processes.

Phylogenetics Study of SalviaL. spp. Collections from the Botanical Garden of Medicinal Plants of Wroclaw Medical University

Systematics of genus Salvia L. is still a field of discussion in taxonomic society. To this day, the position of certain genera from Lamiaceae Martinov. family are considered as autonomic genera or incorporated in the genus Salvia L. as a subgenera (i.e., Perovskia Kar., Rosmarinus L.). Moreover, some species are distinguished only by low-level differences in morphological traits and their geographic occurrence (Drew et al. 2017). In this research, we focused on a molecular analysis of morphologically similar Salvia L. species with special attention paid to Salvia glutinosa L. and Salvia nubicola Wall. ex Sweet. All samples were collected from the Botanical Garden of Medicinal Plants of Wroclaw Medical University (http://www.obrl.umed.wroc.pl/index.html) and the Herbarium of Natural History Museum, Wroclaw University (http://www.muzeum-przyrodnicze.uni.wroc.pl/index.php). Several DNA barcodes, including matK, rbcLa, ITS2 genes, and psbA-trnH intergenic spacer, were used in maximal likelihood and Bayesian inference analyses. All sequences were amplified with Q5 High Fidelity DNA Polymerase (https://www.neb.com/) and universal primers. Amplicons were then sequenced by Sanger sequencing and analysed using the BLAST algorithm (Altschul et al. 1990). Subsequently, sequences were aligned using MAFFT v 7.409 (Katoh et al. 2005) software; poorly aligned sites were objectively eliminated with Gblocks v.0.91b (Talavera and Castresana 2007). ITS2 regions were extracted with ITSx (Bengtsson-Palme et al. 2013) software implemented on PlutoF web workbench (Abarenkov et al. 2010) to obtain ITS2 sequences without 5.8S and 26S fragments on both ends. Substitution models were identified utilizing jModelTest 2 software (Posada 2008) and basing on Bayesian information criterion (BIC) appropriate models were applied for further calculations. For maximum likelihood analyses and Bayesian inference, we implemented RAxML ver. 8.2.10 (Stamatakis et al. 2008) and MrBayes ver. 3.2.2 (Ronquist et al. 2012) respectively. For the reconstruction of evolutionary relationship between study species, different DNA barcode combinations were applied (e.g., matK+ITS2+psbA-trnH and rbcLa+matK+ psbA-trnH). Consensus trees were compared and analysed using TreeGraph ver. 2.14 (Stöver and Müller 2010).

Short Communication: DNA extraction from stored wood of Falcataria moluccana suitable for barcoding analysis

Abstract. Shabrina H, Siregar UJ, Matra DD, Kamiya D, Siregar IZ. 2019. Short Communication: DNA extraction from stored wood of Falcataria moluccana suitable for barcoding analysis. Biodiversitas 20: 1748-1752. Sengon or Falcataria moluccana (Miq.) Barneby & J.W.Grimes is the main crop in community tree plantations in Java, Indonesia, favored because of its fast-growing property and the promising economic return. The wood itself is mostly used for light construction, furniture, plywood, packing materials, and recently as feedstock for bioenergy. Sometimes the wood is used as DNA source for example in the wood identification analysis. However, extracting DNA from woods is considered difficult due particularly to very small quantities of DNA. The objective of this research was to optimize and modify the common CTAB protocols to extract DNA from Sengon wood without liquid nitrogen which sometimes unavailable in some laboratories. The extracted DNA was quantified with nanophotometer and gel visualization and amplified with primer coding psbA-trnH intergenic spacer for testing. The highest concentration of DNA extracted was from 100 mg of wood stored for 24 hours in -30 °C (257.80 ng/µL or 14.18 µg in total) and even the lowest concentration produced by this method able to produce sufficient amount of PCR product for sequencing. Compared with results from 200 mg samples and longer freezing time (72 hours) and extraction using liquid nitrogen, this method considered gave the best results.

Biogeography and phylogenetic relationships of Hyrcanian wild apple using cpDNA and ITS noncoding sequences

The Hyrcanian forest of northern Iran is considered one of the potential centres for the evolution and domestication of the genus Malus (Rosaceae). However, the biogeography, phylogenetic position, and taxonomic status of the Hyrcanian wild apples have never been evaluated. In our study, the nucleotide sequences of the internal transcribed spacer (ITS) and the trnH-psbA intergenic spacer region from 14 natural populations were analysed. Phylogenetic analysis based on the ITS and the Maximum-likelihood (ML) tree showed that all Hyrcanian samples were closely related to M. orientalis and M. asiatica and can be placed within section Malus and series Malus. Furthermore, based on a comparison of ITS2 secondary structures, the Hyrcanian samples were identical to M. orientalis and M. sieversii. Biogeographic scenarios constructed using Statistical Dispersal-Vicariance Analysis (S-DIVA) and the Bayesian Binary Method (BBM) indicated that the ancestor of Malus originated during the Eocene, ∼53 million years ago (Ma), and that China played a vital role in the expansion of the range of the genus. The members of Malus colonized the Hyrcanian region from China during the Miocene, ∼22-10 Ma.

Phylogeography of the palm Syagrus coronata (Martius) Beccari (Arecaceae): distribution in the “Caatinga” and Atlantic forest domains

The palm species Syagrus coronata (Mart.) Becc. shows widespread geographic distribution in the “Caatinga” (seasonally dry tropical forest biome) and Atlantic forest domains (rain forest biome), which suggests that environmental variation in the two domains may have modeled the distribution of S. coronata, generating a distinct phylogeographic scenario. This study assessed the phylogeographic structure of S. coronata in the “Caatinga” and Atlantic forest domains using 181 sequences of the trnH-psbA intergenic spacer from samples spread in ten localities. Genealogical relationships between the haplotypes were estimated using the median-joining method, and spatial analysis of molecular variance was performed using SAMOVA software. The results revealed ten haplotypes, some of which were unique to either the “Caatinga” or the Atlantic forest domain, and analysis of the spatial molecular variance revealed four groups, showing genetic structure (FST = 0.56, P < 0.0001; FCT = 0.51, P < 0.0001), with 51.22% variation between the groups. Remarkably, an inverted sequence of 13 bp was observed in the position 296–308 bp and using the sequence of Cocos nucifera L. as a reference, it was observed that the ancestor type was typical of the “Caatinga” domain and inverted type of the Atlantic forest domain. The results show that S. coronata presents a phylogeographic structuring distributed in the “Caatinga” and Atlantic forest domains. “Caatinga” domain as an ancient refuge and Atlantic forest domain as a regional center of diversification and the Sao Francisco River had an important role in promoting diversification in the “Caatinga” domain.

Pandan (Pandanus sp), Rotan (Calamus sp), and Rengas (Gluta sp) from Kajuik Lake, Riau Province, Indonesia

ABSTRACT Pandan (Pandanus sp), Rotan (Calamus sp), and Rengas (Gluta sp) are the three most important plants growing at Kajuik Lake, Langgam, Riau Province, Indonesia; however, their species names have not been identified. This study aimed to identify their species names using nuclear internal transcribed spacer (ITS) and psbA-trnH intergenic spacer sequences. The method employed was DNA isolation from fresh leaves, PCR using primer pairs of ITS region for Pandanus sp and psbA-trnH intergenic spacer for Calamus sp and Gluta sp, electrophoresis, sequencing, and data analysis using BLASTn program and MEGA software version 6.0. Pandanus tectorius was the only one accession that was similar to Pandanus sp with the identity was 90%, however the query cover was too small, only 39%. On the contrary, Calamus sp showed the highest genetic similarity to Calamus travancoricus, but in fact, both were differed morphologically. There was no database of psbA-trnH intergenic spacer sequence available for species in Gluta. In conclusion, the species names for those plants still could not be determined. It because they might be the identified plants but their sequences databases were not available in large quantities or they were new species which had never been identified and published in public database.