rbcL Sequences Research Articles

Aptychella robusta (Pylaisiadelphaceae) an epiphytic, gemmiferous moss- new to the bryoflora of India

We report the occurrence of an epiphytic, gemmiferous moss, Aptychella robusta (Pylaisiadelphaceae), as an addition to the bryoflora of India. The species was collected from the north-facing mesic site within the Oak forest in the Gairsain area of Chamoli district, Uttarakhand. Earlier, the species was known only from East Asia, Australia, and Central and South America. As A. robusta shows a close morphological resemblance with A. planula, therefore, we amplified and sequenced its rbcL DNA-barcode region. Its comparison with the available rbcL sequences of A. robusta and A. planula on the NCBI database further validated the identity of the moss as A. robusta. The key morphological characteristics of this taxon and the rbcL amplicon have been given for future reference.

Of sea, rivers and symbiosis: Diversity, systematics, biogeography and evolution of the deeply diverging florideophycean order Hildenbrandiales (Rhodophyta)

The Hildenbrandiales, a typically saxicolous red algal order, is an early diverging florideophycean group with global significance in marine and freshwater ecosystems across diverse temperature zones. To comprehensively elucidate the diversity, phylogeny, biogeography, and evolution of this order, we conducted a thorough re-examination employing molecular data derived from nearly 700 specimens. Employing a species delimitation method, we identified Evolutionary Species Units (ESUs) within the Hildenbrandiales aiming to enhance our understanding of species diversity and generate the first time-calibrated tree and ancestral area reconstruction for this order. Mitochondrial cox1 and chloroplast rbcL markers were used to infer species boundaries, and subsequent phylogenetic reconstructions involved concatenated sequences of cox1, rbcL, and 18S rDNA. Time calibration of the resulting phylogenetic tree used a fossil record from a Triassic purportedly freshwater Hildenbrandia species and three secondary time points from the literature. Our species delimitation analysis revealed an astounding 97 distinct ESUs, quintupling the known diversity within this order. Our time-calibration analysis placed the origin of Hildenbrandiales (crown age) in the Ediacaran period, with freshwater species emerging as a monophyletic group during the later Permian to early Triassic. Phylogenetic reconstructions identified seven major clades, experiencing early diversification during the Silurian to Carboniferous period. Two major evolutionary events—colonization of freshwater habitats and obligate systemic symbiosis with a marine fungus—marked this order, leading to significant morphological alterations without a commensurate increase in species diversification. Despite the remarkable newly discovered diversity, the extant taxon diversity appears relatively constrained when viewed against an evolutionary timeline spanning over 800 million years. This limitation may stem from restricted geographic sampling or the prevalence of asexual reproduction. However, species richness estimation and rarefaction analyses suggest a substantially larger diversity yet to be uncovered—potentially four times greater. These findings drastically reshape our understanding of the deeply diverging florideophycean order Hildenbrandiales species diversity, and contribute valuable insights into this order's evolutionary history and ecological adaptations. Supported by phylogenetic, ecological and morphological evidence, we established the genus Riverina gen. nov. to accommodate freshwater species of Hildenbrandiales, which form a monophyletic clade in our analyses. This marks the first step toward refining the taxonomy of the Hildenbrandiales, an order demanding thorough revisions, notably with the creation of several genera to address the polyphyletic status of Hildenbrandia. However, the limited diagnostic features pose a challenge, necessitating a fresh approach to defining genera. A potential solution lies in embracing a molecular systematic perspective, which can offer precise delineations of taxonomic boundaries.

Old World species of Trichanthecium (Poaceae: Panicoideae: Paniceae) including a new species from the Democratic Republic of Congo

Studies of African material of the tribe Paniceae (Poaceae) resulted in the transfer of 12 species from Panicum, the description of a new species, and the conclusion that the genus Trichanthecium now consists of 52 species. The new Trichanthecium delimitation was confirmed by phylogenetic studies based on chloroplast ndhF and rbcL sequences. Morphological and anatomical characters are discussed within their pylogenetic context. A new species is described: T. marunguensis. Lectotypes are designated for Panicum aequinerve, P. calvum, P. chionachne, P. hochstetteri var. glaberrimum, P. inaequilatum, P. lepidum, P. mokaense, and P. wiehei. The following new combinations are proposed: T. acrotrichum, T. aequinerve, T. calvum, T. chionachne, T. delicatulum, T. eickii, T. hochstetteri, T. inaequilatum, T. perrieri, T. pusillum, T. striatissimum, and T. wiehei. Full synonymies, descriptions, distribution and habitat information, citation of all specimens and a key to the Old World species of the genus are provided; seven species are illustrated.

Two new species of Dictyota (Dictyotales, Phaeophyceae), D. dimorphosa sp. nov. and D. recumbens sp. nov., based on morphological and molecular investigations

ABSTRACT The brown algal genus Dictyota (Dictyotales, Phaeophyceae) is widespread, particularly in warm temperate and tropical waters, and plays a vital role in coastal ecosystems. This study reassessed the taxonomy of Dictyota, focusing on specimens from Myanmar and Japan, using a combination of morphological and molecular analyses. Molecular phylogenetic analyses using concatenated DNA sequences of rbcL, psaA, psbA, cox1 and cox3 genes revealed that Dictyota specimens from Myanmar, Japan, and other regions, belong to nine different clades, each representing a different species. Based on morphological and molecular data, seven out of these nine clades are identified as known species, while the remaining two clades are described as new species D. recumbens sp. nov. (=D. sp. A) and D. dimorphosa sp. nov. (=D. sp. B). Dictyota recumbens is characterized by the recumbent thallus, with transverse yellowish-green iridescent lines, and the formation of solitary spores irregularly distributed over the thallus. Dictyota dimorphosa is characterized by two morphological forms: decumbent and erect thalli, with bluish iridescence mainly in the decumbent thallus, and the formation of spores in oval ring shapes distributed over both thallus surfaces. Dictyota recumbens was typically found in tropical regions, while D. dimorphosa appears to be a cool-warm temperate species, distributed from northeastern Hokkaido to southern Kyushu with an extension along South Korea. Our molecular phylogenetic analyses have revealed the existence of several species whose names appeared in multiple clades, suggesting the potential for future increases in species diversity and the presence of cryptic species within certain groups.

Phymatolithopsis acervata comb. nov., and P. roseola sp. nov. (Hapalidiales, Rhodophyta) from South Africa with a comment on infraordinal classification

Abstract Phymatolithon acervatum is an encrusting, non-geniculate coralline alga from South Africa. While morpho-anatomical details of the species are well documented, no genetic analyses have been performed on P. acervatum. Here, we analyzed rbcL and psbA gene sequences, and they showed that two species were passing under the name Phymatolithon acervatum in South Africa. A partial rbcL sequence from the lectotype of Lithothamnion acervatum, basionym of P. acervatum, showed to which species the specific epithet applies, and furthermore that both species belong in Phymatolithopsis. We transfer Phymatolithon acervatum to Phymatolithopsis, as Phymatolithopsis acervata comb. nov., and propose Phymatolithopsis roseola sp. nov. These species overlap in morpho-anatomy, habitat and distribution and can only be distinguished based on DNA sequences. Previous misinterpretations of conceptacle development are corrected for species of Phymatolithopsis. The main character used to define the family Mesophyllumaceae is shown to have arisen several times in independent lineages in the order Hapalidiales. At present, only one family, Hapalidiaceae, thus merits recognition in the Hapalidiales.

Reappraisal of the genus Visia (Batrachospermales, Rhodophyta) and proposal for a new genus and species

ABSTRACT The genus Visia is reappraised based on new morphological and molecular data (rbcL and COI-5P sequences), from recently collected and previously studied materials, resulting in the need for several taxonomic changes. Based on our phylogeny and sequence divergence, the genus Visioidea gen. nov. is proposed for two samples from Brazil, forming a distinct clade that is morphologically undistinguishable from the generitype of Visia (V. cayennensis). Two lineages of Visia with high genetic divergences from other species are proposed as new species: 1) V. verviridis sp. nov. from Australia; 2) V. amazonensis sp. nov. from Brazil. Although genetically distinct, both species are morphologically very similar to V. cayennensis. A new record of V. cylindrocellularis was found in Australia, expanding the geographic distribution and the morphometric ranges for this species. The number of species currently recognized in Visia is now six, four with DNA sequence data and two identified based solely on morphology (V. longiarticulata and V. turgida). Cryptic genera within the order Batrachospermales were not recognised until recently with the proposals of Volatus, morphologically indistinct from Kumanoa, and Macrosporophycos undistinguishable from Montagnia and here a third cryptic genus Visioidea is described.

Morphological analysis, agronomic characteristics, species identification, and phylogenetic analysis of Pseuderanthemum sp. using the rbcLgene

The study aimed to evaluate the agronomic characteristics of eleven samples of Pseuderanthemum sp. variety based on the results of the morphological survey on stem height, root length, leaves, combining molecular biology methods to determine their genetic relationships by building a phylogenetic tree based on the plastid gene rbcL sequence. Initial results of an evaluation of the genetic diversity of eleven Pseuderanthemum sp. samples showed that there were differences in stem length, leaf width to root length. Based on the analysis results from the pedigree tree, the tree samples with similar survival indexes and genetically close to each other were classified into two main groups. Based on molecular data, eleven varieties of Pseuderanthemum sp. in the study have been identified, all belonging to the species Pseuderanthemum sp. have a high similarity coefficient above 98% and 100% coverage. Although the geographical location of the samples is variable, it is necessary to investigate other gene sequences related to morphological and biochemical characteristics, such as bioactive compounds in leaves and fruits to have more accurate conclusions about this species.

Chloroidium phycobionts (Watanabeales, Trebouxiophyceae) partner with lecanoralean mycobionts in foliicolous lichen communities of Tenerife (Canary Islands) and Navarra (Iberian Peninsula), Spain

Abstract While the diversity of foliicolous lichen-forming fungi has been explored in substantial depth, relatively little attention has been paid to their algal symbionts. We studied the unicellular green phycobionts of the lecanoralean lichens Bacidina (Ramalinaceae), Byssoloma, Fellhanera and Tapellaria (Pilocarpaceae) and graphidalean Gyalectidium (Gomphillaceae) from two extratropical foliicolous communities in continental Spain and the Canary Islands. We examined the pyrenoids of algal symbionts within thalli using TEM, and obtained several algal nrSSU and rbcL sequences from whole thalli, and also from cultures isolated from some of these lichens. Pyrenoid structure and molecular sequence data provided support for recognizing Chloroidium (Watanabeales, Trebouxiophyceae) as phycobiont in thalli of Byssoloma subdiscordans and Fellhanera bouteillei (Pilocarpaceae) in both communities. Bacidina apiahica (Ramalinaceae) and Tapellaria epiphylla (Pilocarpaceae) likewise appeared to partner with Chloroidium based on the presence of the same pyrenoid type, although we were able to obtain a phycobiont sequence only from a culture isolate of the latter. These results contrast with those obtained previously from a foliicolous lichen community in southern Florida, which revealed only strains of Heveochlorella (Jaagichlorella) as phycobiont of foliicolous Pilocarpaceae and Gomphillaceae. On the other hand, the pyrenoid we observed in the phycobionts associated with Gyalectidium setiferum and G. minus corresponded to that of Heveochlorella (Jaagichlorella). However, the poor quality of the phycobiont sequence data obtained from G. minus, probably due to the presence of epibiontic algae, could not provide additional perspective on the pyrenoid structure observations. Nonetheless, clear differences in pyrenoid ultrastructure can allow Chloroidium and Heveochlorella phycobionts to be distinguished from each other in TEM. Our results indicate a greater diversity of unicellular green-algal symbionts in foliicolous communities from Spain than previously observed in other geographical areas, and suggest that further studies focused on symbiont pairing in these communities might reveal distinctive and varied patterns of phycobiont preference.

Genetic analysis of Ulva (Ulvaceae, Chlorophyta) type specimens resolves northeast Pacific blade-forming species

Abstract Misapplication of Ulva epithets in GenBank has led to confusion in the scientific literature and community. To solve some of the problems, targeted DNA sequencing of plastid encoded rbcL gene amplicons or high-throughput sequencing was performed on all blade-forming Ulva type specimens from the northeast Pacific. Recently collected specimens from at or near type localities were also analyzed for some taxa. Based on these genetic analyses, we confirmed currently recognized species: U. californica, with U. angusta and U. scagelii as heterotypic synonyms, U. stenophylla, U. taeniata, and U. tanneri. Ulva dactylifera, currently considered a synonym of U. taeniata based on morpho-anatomy, is recognized as a distinct species, as is U. expansa whose type specimen was sequenced in 2018. All but two of the ITS, rbcL and tufA sequences in GenBank that were labeled U. californica were correctly named, in contrast to U. taeniata, for which only one of 14 sequences was correctly labeled. These results show that DNA sequencing of Ulva type specimens is essential for the correct application of names.

Taxonomic study of the polyphyletic Dudresnaya (Dumontiaceae, Florideophyceae) with descriptions of Dudresnaya ryukyuensis sp. nov. and two new genera, Himehibirhodia and Nudresdaya

SUMMARYThe red algal genus Dudresnaya (Dumontiaceae, Gigartinales) has traditionally been a morphologically well‐defined taxon, but its molecular phylogeny has rarely been studied. To examine the phylogenetic relationships among Dudresnaya species, we generated new partial sequences of mitochondrial cox1, chloroplast rbcL and nuclear 28S rRNA genes from an undescribed Dudresnaya species from Okinawa Island, Japan, alongside five additional described species. Our phylogenetic analyses show that Dudresnaya is genetically diverse and polyphyletic. Based on molecular phylogeny and morphological data, we describe the Okinawan Dudresnaya as a new species, Dudresnaya ryukyuensis, and transferred Dudresnaya minima and Dudresnaya littleri, which were phylogenetically and morphologically distinct from the genuine Dudresnaya, to the new genera Himehibirhodia and Nudresdaya, respectively. Our phylogenetic analyses also showed that the Dumontiaceae is not a monophyletic group including the Gainiaceae and Rhizophyllidaceae (DGR complex). Considering that the DGR complex exhibits female reproductive structures and their post‐fertilization development that are similar to each other, the DGR complex appears to be recognized as the Dumontiaceae sensu lato.

Updating the Ulvaceae in the green seaweeds of Britain and Ireland

Abstract The 2007 flora “Green Seaweeds of Britain and Ireland” did not present the molecular data underpinning the Ulvaceae treatment, mostly ITS sequences. Subsequently, names have changed as type material of Ulva species is sequenced and intensive sampling with DNA barcoding adds new European species. To update the Ulvaceae, we systematically sampled from 2007 to 2021, identifying specimens using various molecular markers alongside DNA from type material of four species. We show here that Ulva gigantea, based on rbcL, tufA and ITS sequences of its holotype, is assigned to Ulva compressa, as is the lectotype of Ulva curvata. Ulva gigantea sensu GenBank and Ulva pseudocurvata are conspecific. The correct name is U. pseudocurvata based on rbcL sequences of the lectotype. Two species of monostromatic Ulvaceae were included in the 2007 flora, but we show that both of them and all earlier British monostromatic collections represent Ulvaria splendens, a species originally described from Alaska. Analysis of two rbcL amplicons of the Ulva sordida lectotype shows that it is conspecific with Ulvaria splendens. Our first genuine collections of Ulvaria obscura from SW England and SW Wales correspond to topotype material from the Bay of Biscay, recent samples from Galicia and unpublished tufA sequences from Britanny.

Phylogenetic affiliation of Pedinomonas noctilucae and green Noctiluca scintillans nutritional dynamics in the Gulf of Mannar, Southeastern Arabian Sea

The present investigation focused on studying the phylogenetic position of the green Noctiluca endosymbiont, Pedinomonas noctilucae, collected from the Gulf of Mannar, India. In this study, we re-examined the evolutionary position of this endosymbiotic algae using rbcL sequences. The phylogenetic analysis revealed that P. noctilucae is distantly related to the Pedinomonas species, and formed a monophyletic clade with Marsupiomandaceae. Based on the phylogenetic association of endosymbiont with Maruspiomonadales it was concluded that the endosymbiont belongs to an independent genus within the family Marsupiomonadaceae. At the site of the bloom, Noctiluca scintillans was found to exhibit a dense monospecific proliferation, with an average cell density of 27.l88 × 103 cells L−1. The investigation revealed that the green Noctiluca during its senescent phase primarily relied on autotrophic nutrition, which was confirmed by the presence of a high number of trophonts, vegetatively reproducing cells (1.45 × 103 cells L−1) and the absence of food vacuoles.

Striking differences in frost hardiness and inability to cold acclimate in two Mougeotia species (Zygnematophyceae) from alpine and lowland habitats

AbstractZygnematophyceae, a class of freshwater green algae, exhibit distinctive seasonal dynamics. The increasing frequency of cold snaps during the growing season might challenge the persistence of some populations. The present study explored the frost hardiness of two Mougeotia species isolated from different elevations and habitats. Additionally, a phylogenetic (rbcL sequence), ultrastructural and physiological characterization was performed. Both species, grown under standard culture conditions and cold acclimated cultures (+4°C), were exposed to freezing temperatures down to −9°C. Furthermore, ultrastructural‐, hydrogen peroxide (H2O2)‐ and photosynthetic pigment analysis were performed on cells exposed to −2°C, with and without induced ice nucleation. The alpine M. disjuncta showed a higher frost hardiness (LT50 = −5.8°C), whereas the lowland M. scalaris was susceptible to ice. However, frost hardiness did not improve after cold acclimation in either species but rather decreased significantly in M. disjuncta (LT50 = −4.7°C). Despite darkness, prolonged sub‐zero temperatures or freezing induced the activation of the xanthophyll (VAZ) cycle in M. scalaris. Our results demonstrate that frost hardiness varies within the genus Mougeotia and that the VAZ cycle can be activated in the dark under subzero temperature‐ and freezing stress but does not necessarily increase frost hardiness. As highly frost hardy cell types are usually formed at the end of the growing season, the ability of young cells to survive ice formation in the upper subzero temperature range represents a crucial survival strategy in populations exposed to late spring frosts.

Morphology and phylogeny of Nitzschianandorii sp. nov. (Bacillariophyceae), a new small-celled lanceolate species from a post-mining reservoir.

Post-mining reservoirs are distinguished by characteristic environmental conditions where specific diatom communities can be observed. Reservoirs created as a part of the reclamation plan after human mining activities are marked by unique chemical and physical water parameters. In the course of research on the diatoms from Bogdałów reservoir, we examined the taxonomic and morphological diversity of Nitzschia taxa from the section Lanceolatae occurring in a post-mining lignite reservoir. Our study describes a new species of Nitzschia from a post-mining reservoir, Nitzschianandorii Olszyński, Zakrzewski & Żelazna-Wieczorek, sp. nov. Morphometry and morphology analyses of new species were performed with light and scanning electron microscopy. Chloroplast morphology analysis was conducted with differential interference contrast microscopy and confocal laser scanning microscopy. Molecular data from SSU 18S, rbcL and psbC sequences were obtained from cultures of this taxon. Differential diagnosis of Nitzschianandorii Olszyński, Zakrzewski & Żelazna-Wieczorek, sp. nov. with co-occurring taxa: N.lacuum and N.alpinobacillum was performed using morphological traits and nMDS analysis of the valves' morphometry.

Molecular classification of Barbeyaceae (Barbeya oleoides Schweinf.) using four different DNA barcodes

Despite many efforts to determine the phylogenetic relationship between Barbeyaceae and other families of Rosales, the sister relationship of this family has remained unclear. Barbeya oleoides, which is currently the only species in the family Barbeyaceae, and native to Somalia, Ethiopia, and the Arabian Peninsula were collected from Wadi Turbah Zahran northwestern of Al-Baha City, southwestern Saudi Arabia (20°14’N, 41°15’E). To study the sister relationship of Barbeyaceae and the other families of Rosales, the complete chloroplast sequences were used for phylogenetic analysis. In addition, four standard DNA barcodes (the internal transcribed spacer 2 (ITS2), ribulose 1,5-biphosphate carboxylase (rbcL), maturase K (matk), the intergenic spacer region (trnH-psbA)) were used to test for their quality in identifying phylogenetic relationship of the studied families. Sequence analysis of the complete chloroplast sequences showed that Rosales clade has two subclades and clearly discriminated all families within this order. This is the first report of a partial ITS2 locus sequence in B. oleoides. The partial ITS2, rbcL, matk, and gene sequences discriminate B. oleoides of the Barbeyaceae family from the closely related plant families: Cannabaceae, Rosaceae, Rhamnaceae, Urticaceae, Moraceae, Elaeagnaceae, Dirachmaceae, and Ulmaceae. In contrast, the partial trnH-psbA sequence of B. oleoides did not show any homology to the available DNA sequence of plant families in GenBank, suggesting that it is more suitable as DNA barcode for variations within one species.

Maximising environmental pressure-response relationship signals from diatom-based metabarcoding in rivers

DNA metabarcoding has been performed on a large number of river phytobenthos samples collected from the UK, using rbcL primers optimised for diatoms. Within this dataset the composition of non-diatom sequence reads was studied and the effect of including these in models for evaluating the nutrient gradient was assessed. Whilst many non-diatom taxonomic groups were detected, few contained the full diversity expected in riverine environments. This may be due to the performance of the current primers in characterising the wider phytobenthic community and influenced by the sampling method employed, as both were developed specifically for diatoms. Nevertheless, the study identified considerable diversity in some groups, e.g. Eustigmatophyceae and a wider distribution than previously thought for freshwater Phaeophyceae. These results offer a strong case for the benefits of metabarcoding for expanding knowledge of aquatic biodiversity in the UK and elsewhere.Many of the ASVs associated with non-diatoms showed significant pressure responses; however, models that included non-diatoms had similar predictive strength to those based on diatoms alone. Whilst limitations of the primers for assessing non-diatoms may play a role in explaining these results, the diatoms provide a strong signal along the nutrient gradient and other algae, therefore, add little unique information.We recommend that future developments should use ASVs to calculate metrics, with links to reference databases made as a final step to generate lists of taxa to support interpretation. Any further exploration of the potential of non-diatoms would benefit from access to a well-curated reference database, similar to diat.barcode. Such a database does not yet exist, and we caution against the indiscriminate use of NCBI GenBank as a taxonomic resource as many rbcL sequences deposited have not been curated.

Development of specific barcodes for identification of Aloe species based on chloroplast DNA barcoding

DNA barcoding is a widely used taxonomic tool for correctly identifying plant species. The chloroplast matK and rbcL are some of the most extensively studied variable coding genes of angiosperms and have been recommended as a universal barcode for terrestrial plants. Genus Aloe contains around 500 species, and most Aloe species have extensive traditional medicinal usage worldwide, viz., A. arborescens, A. ferox, A. maculate, and A. vera. This study evaluated the two chloroplast DNA barcodes (matK and rbcL) to develop a theoretical base for species identification and germplasm conservation of Aloe species. The Maximum Parsimony analysis was conducted to study the evolutionary relatedness of Aloe sequences using matK and rbcL sequences obtained from the NCBI nucleotide database. The results revealed that 49 Aloe accessions were segregated into four major clades using matK phylogenetic analysis. Likewise, the 29 Aloe accessions were allocated into two clades based on the rbcL phylogeny. Additionally, specific barcodes have been developed for the following species: A. vera, A. boylei, A. ciliaris, A. karasbergensis, and A. sinkatana based on SNP (Single Nucleotide Polymorphism) site analysis of barcode sequences and the corresponding DNA QR (quick response) code of each sequence was generated. This study provides a unique method for species identification of the genus Aloe and indicates the possibility of correctly identifying, discriminating, and documenting the Aloe species.

Molecular Phylogenetics and Molecular Dating of Arecaceae In Bangladesh Inferred from Matk and Rbcl Genes

A molecular phylogenetic investigation was undertaken for 30 species belonging to 15 genera of the palm family Arecaceae in Bangladesh to infer evolutionary relationships and molecular dating utilizing plastid-based matK and rbcL genes through multifaceted-algorithm driven approaches with Neighbor-Joining, Maximum-Likelihood, and Bayesian Inference methods. The study revealed that matK has better species discrimination efficiency than rbcL gene due to its highly variable nature. Transition/transversion bias test corroborated this finding as matK showed higher bias (2.632) than rbcL (2.235). Nucleotide substitution patterns were visualized via HYPERMUT program, which unveiled higher variability in matK and lower variability in rbcL alignment. Phylogenetic trees constructed with matK revealed monophyletic nature of origin for all the three subfamilies, viz. Arecoideae, Coryphoideae and Calamoideae, while rbcL trees exhibited polyphyly for Coryphoideae and monophyly for Arecoideae and Calamoideae. All the nine tribes belonging to three subfamilies demonstrated monophyletic nature in matK trees. Bootstrap support and Bayesian posterior probability were found to be higher in matK topologies than that of rbcL. The molecular clock test unraveled an equal evolutionary rate for matK and unequal rate for rbcL sequences. Molecular dating approach unveiled Calamoideae to be the most ancient subfamily (65.75 MYA) among the three subfamilies that originated during the Late Cretaceous period in the Mesozoic era, whereas Coryphoideae and Arecoideae were found to have originated in the Cenozoic era. Bangladesh J. Plant Taxon. 30(2): 213-232, 2023 (December)

DNA barcoding of an endangered plant species, salvadora oleoides

Salvadora oleoides Decne is an endangered plant species. This species is important as the plant is used for multiple purposes. Natives use the plants for their medicinal needs in treating rheumatic pains and coughs, and many others. This species is threatened by over-exploitation and deforestation or illegal logging, as well as its very slow growth rate, so the accurate identification of this species is important. Knowledge about threats or even distribution is limited, making identifying species more difficult. Therefore, this research aimed at DNA barcoding of endangered species Salvadora oleoides to assess their potential at the meaningful taxonomic level. This study aims to amplify the rbcL and matK genes in Salvadora oleoides collected from the different regions of Pakistan. The plant DNA isolation was carried out by the CTAB method. PCR analysis was performed, which showed amplification of both rbcL and matK regions. The amplified products were purified and sent for sequencing. The results revealed a 100% sequencing rate for rbcL; however, the matK amplified product was not sequenced due to the high heterozygosity in this region, which might result in polymerase slippage during the Sanger sequencing method. The rbcL sequence was checked for homology by BLASTn and then submitted to NCBI for GenBank accession. The accession number assigned to Salvadora oleoides was OP046316 for rbcL. Keywords: DNA Barcoding, Medicinal Plants, matk, rbcl, BLAST, Species Identification.

Effectiveness of DNA barcoding, SCOT markers and phytochemical characterization in biodiversity assessment of some Zea mays hybrids

Because of the rapid fluctuation in world's climate, politics, and economy, maize outputs must be increased to meet food security demands. Due to the importance of the genetic materials under study as a source of quality traits and genetic variability, this study was designed to examine the effectiveness and strength of assessing biodiversity in some maize hybrids using barcodes and molecular SCoTs markers in addition to phytochemical characterization. Using matK and rbcL sequences, pairwise distances were obtained and assessed. The hybrids under study were distributed into different groups based on the matK and rbcL sequences. The 12 SCoT primers amplified 155 amplicons with an average of 12.92 bands per primer, of which 63 monomorphic bands (MB) (40.65 %) and 92 polymorphic band (PB) (59.35 %). Additionally, the number of unique bands was six bands. The SCoT-6 primer produced three bands individually, while the primers SCoT-2, SCoT-3, SCoT-4, and SCoT-7 produced one band each. Hybrids S.C. 10, S.C. 128, S.C. 130, S.C. 131, S.C. 132 and TWC. 324 were more influenced by some primary constituents and bioactive secondary compounds such as moisture content (%), crude fiber, total carbohydrates, total sugars, soluble carbohydrates, non-reducing sugars, phenolics compounds, and tannins. The hybrids S.C. 168, S.C. 176, S.C. 178, TWC. 360, TWC. 368 and TWC. 321 were more influenced by the SCoT molecular markers. The results of genetic variation between the studied hybrids could be considered a potentially source of genetic information in developing future breeding programs.