rbcL Gene Research Articles

Ulva L. (Ulvales, Chlorophyta) from Manawatāwhi/ Three Kings Islands, New Zealand: Ulva piritoka Ngāti Kuri, Heesch & W.A.Nelson, sp. nov. and Records of Two Nonnative Species, U. compressa and U. rigida

Manawatāwhi/Three Kings Islands lie to the north of the North Island of New Zealand. Manawatāwhi is part of the rohe (territory) of Ngāti Kuri, with the islands having spiritual, cultural, political and customary significance. This group of small islands has one of the most pristine coastlines in New Zealand, with no human-mediated impacts from current land use, no permanent anchorage points, and landing on the islands is prohibited. The islands harbour a rich marine biota with a number of endemic species. A recent collection trip and molecular genetic studies using the rbcL marker revealed the presence of three entities of the genus Ulva L. (Ulvales, Chlorophyta) on the islands. One of these we describe herein as Ulva piritoka Ngāti Kuri, Heesch & W.A.Nelson, sp. nov. While it is genetically close to U. adhaerens Matsumoto & Shimada from Japan, differences in morphology, habitat and rbcL gene region, including the presence of a large group II intron in U. adhaerens, which was lacking in the Manawatāwhi specimen, led us to conclude U. piritoka sp. nov. belongs to a distinct species. We also confirmed the presence of Ulva compressa L. and Ulva rigida C.Agardh, both of which, although present on the New Zealand mainland, were originally described from Europe and are considered introductions to the New Zealand archipelago. These are the first reported marine non-indigenous species (NIS) recorded from Manawatāwhi/Three Kings Islands, and the implications of their presence are discussed.

Description and phylogenetic position of three new species of Stauroneis Ehrenberg (Bacillariophyceae: Stauroneidaceae) from the Indian Subcontinent

ABSTRACT Stauroneis Ehrenberg is a species-rich genus that is common and widespread with some variation in frustule morphology. Recently, studies have explored the molecular diversity of the Stauroneis from the temperate region, but there are no molecular data for tropical species. In the present study, four species of Stauroneis were investigated using morphological and molecular data. Morphological characters were analysed using light microscopy (LM) and scanning electron microscopy (SEM), and the features of each taxon were compared with similar species within the genus. For the molecular analyses, Stauroneis strains were isolated, cultured, DNA was extracted, and sequences from rbcL and 18S genes were analysed to determine phylogenetic relationships. Our analysis describes three new species and records one previously known species. The new species are: (1) Stauroneis lateritica Wadmare, Kociolek & B.Karthick, characterized by small frustules with elliptical lanceolate valves and short subrostrate apices, broad central bow-tie shaped stauros, and few moderate striae centrally becoming strongly radiate towards the poles; (2) Stauroneis sholaii Wadmare, Kociolek & B.Karthick, which has broad lanceolate valves with short rectangular stauros and radiate striae; and (3) Stauroneis bartii Wadmare, Kociolek & B.Karthick, which has large, lanceolate valves with rounded apices, a stauros with 4–7 shortened striae and radiate striae. Ultrastructure and molecular data from India for the previously known, widespread species Stauroneis gracilis Ehrenberg are presented. The combined morphological and molecular approach supports recognition of S. lateritica, S. sholaii and S. bartii as new species. This study is the first-ever attempt at molecular species discovery of diatoms from the Indian subcontinent.

Succinic acid inhibits photosynthesis of Microcystis aeruginosa via damaging PSII oxygen-evolving complex and reaction center.

To elucidate the mechanism of succinic acid (SA) inhibition of Microcystis aeruginosa, the chlorophyll fluorescence transients, photosynthesis, photosynthetic electron transport activity, and gene expression of M. aeruginosa were evaluated under various doses of SA. The results demonstrated that, after treatment with 60 mg L-1 SA for 1 h, the chlorophyll fluorescence transients and related parameters changed significantly, indicating that the function and structure of photosynthetic apparatuses of Microcystis were seriously damaged. The initial quantum efficiency α, maximum net photosynthetic rate Pnmax, dark respiration rate Rd, and gross photosynthetic rate decreased to 57%, 49%, 49%, and 46%, respectively, relative to the control. Furthermore, photosystem II (PSII) activity (H2O→p-BQ) and the electron transport activity of H2O→MV and DPC→MV significantly decreased. Real-time PCR analysis revealed that, following incubation with 60 mg L-1 SA for 24 h, the expression level of core protein genes (psbA, psaB, psbD, and psbO) of the photosynthesis centers photosystem I (PSI) and PSII decreased significantly. However, the transcription of gene nblA encoding phycobilisome degradation protein was elevated. The downregulation of the rbcL gene, which encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), resulted in the suppression of CO2 fixation and assimilation. High concentration (60 mg L-1) of SA resulted in damage to oxygen-evolving complex (OEC) and reaction center of PSII, blocking photosynthetic electron transport, thereby lowering the rate photosynthesis and inhibiting the growth of Microcystis. We concluded that inhibition of photosynthesis is an important mechanism of SA inhibition in M. aeruginosa.

Enhancement of Lipid Productivity and Self-flocculation by Cocultivating Monoraphidium sp. FXY-10 and Heveochlorella sp. Yu Under Mixotrophic Mode.

To maintain high microalgae lipid productivity and flocculation efficiency simultaneously and reduce the production cost of microalgae lipids, Monoraphidium sp. FXY-10 with high lipid-producing capacity and Heveochlorella sp. Yu with strong self-flocculation ability were cocultivated and studied. Cocultivated microalgae lipid productivity and flocculation efficiency were increased to 203.8 mg L-1 day-1 and 70.55%, respectively, which is potentially related to the excessive competitive depletion of nitrogen sources and the upregulation of correlative key genes in lipid anabolic metabolism. Under cocultivation conditions, microalgae cells could enter the stationary phase 2 days earlier than that under monocultivation conditions, thus reducing the culture time. Relative expression of the accD, ME, and rbcL genes was upregulated to varying degrees, and the enzyme activities of ACCase, ME, and RuBisCO were also significantly increased compared with those in monocultivation. Moreover, fatty acid composition showed that microalgae lipids in cocultivation exhibited potential as a feedstock for biodiesel.

Plant DNA Barcode as a Tool for Root Identification in Hypogea: The Case of the Etruscan Tombs of Tarquinia (Central Italy).

Roots can produce mechanical and chemical alterations to building structures, especially in the case of underground historical artifacts. In archaeological sites, where vegetation plays the dual role of naturalistic relevance and potential threat, trees and bushes are under supervision. No customized measures can be taken against herbaceous plants lacking fast and reliable root identification methods that are useful to assess their dangerousness. In this study, we aimed to test the efficacy of DNA barcoding in identifying plant rootlets threatening the Etruscan tombs of the Necropolis of Tarquinia. As DNA barcode markers, we selected two sections of the genes rbcL and matK, the nuclear ribosomal internal transcribed spacer (nrITS), and the intergenic spacer psbA-trnH. All fourteen root samples were successfully sequenced and identified at species (92.9%) and genus level (7.01%) by GenBank matching and reference dataset implementation. Some eudicotyledons with taproots, such as Echium italicum L., Foeniculum vulgare Mill., and Reseda lutea L. subsp. lutea, showed a certain recurrence. Further investigations are needed to confirm this promising result, increasing the number of roots and enlarging the reference dataset with attention to meso-Mediterranean perennial herbaceous species. The finding of herbaceous plants roots at more than 3 m deep confirms their potential risk and underlines the importance of vegetation planning, monitoring, and management on archaeological sites.

Molecular identification and phylogenetic analysis of Chlorella isolates from Indonesia using rbcL gene

Identifying the newly isolated species is crucial to establishing a reliable algal database with successful commercial applications for different biotechnological applications. Morphological identification does not give sufficient description, especially for tiny unicellular microalgae. The rbcL gene encodes the large unit of ribulose-1, 5-bisphosphate carboxylase /oxygenase (Rubisco) has been widely known for barcoding in plants and developed for microalgae molecular identification. In this study, we examined the local strains of green microalgae from Indonesia using the rbcL partial gene sequence to identify the strains. Green microalgae isolates originated from Yogyakarta, Serayu, Gondol, Ancol, Cilegon, and Teluk Jakarta were cultured in f/2 media and harvested for DNA extraction. The DNA extracted was proceeded to PCR using 1AB_rbcL primer pair to amplify the sequences of rbcL gene with target band located at 582 bp, followed by the sequencing of the PCR product was conducted. Molecular identification of local green microalgae isolates was successfully carried out using primers 1AB_rbcL with a genetic similarity of 99% toward identified species in the NCBI database. Among six isolates, TJ, G, S, C, and A isolates were identified as C. pyrenoidosa. Only CP isolate from Yogyakarta identified as C. sorokiniana. Nannochloropsis gaditana rbcL sequence was selected as an outgroup. The phylogenetic analysis indicated that the five isolates of Chlorella belong to one clade and clearly distinguished from C. sorokiniana isolate from Yogyakarta.

Melanothamnus macaronesicus Rodríguez-Buján & Díaz-Tapia, sp. nov. (Rhodomelaceae, Rhodophyta): A New Turf-Forming Species from the Azores and the Canary Islands

The use of molecular tools in red algal diversity surveys often reveals the existence of undescribed species. Here, we report a new Macaronesian turf-forming red alga in the otherwise mostly Pacific genus Melanothamnus Bornet & Falkenberg. This new taxon, Melanothamnus macaronesicus Rodriguez-Bujan & Diaz-Tapia, sp. nov. is described based on morphological and molecular (rbcL gene) evidence. Morphologically, it differs from other Melanothamnus taxa by a combination of characters that includes its decumbent habit with an extensive system of prostrate axes, small size (≤2.5 cm), four ecorticate pericentral cells, unbranched to once-branched trichoblasts and spermatangial branches formed on the first dichotomy of trichoblasts that have a sterile apical cell. Molecularly, this new species differs from its congeners by sequence divergence ≥3.3% in the rbcL gene. Current known distribution of M. macaronesicus Rodriguez-Bujan & Diaz-Tapia, sp. nov. is restricted to Macaronesia, suggesting that it could be endemic to this archipelago. Our findings suggest that endemicity among red algae might be more common than previously thought in this bioregion, particularly among easily overlooked turf-forming species.

DNA sequencing of type material and newly collected specimens reveals two heterotypic synonyms for Harveylithon munitum (Metagoniolithoideae, Corallinales, Rhodophyta) and three new species.

Nongeniculate coralline algae are difficult to identify based solely on morpho-anatomy. To address the systematics of several taxonomically challenging taxa, we analyzed DNA sequences of a short portion (118-296 base pairs) of the 3' end of the rbcL gene from three type specimens. The analyses revealed that Harveylithon munitum (basionym: Lithophyllum munitum), described in 1906 from Cave Cays, Exuma Chain, Bahamas, is conspecific with both Goniolithon accretum and Goniolithon affine, described in 1906 from Sand Key, Florida and in 1907 from Culebra Island, Puerto Rico, respectively. Lithophyllum munitum and G.accretum were described in the same 1906 publication and have equal priority. We have selected the currently accepted and most commonly used name H.munitum to apply to this entity. Comparative analyses of rbcL, psbA, UPA, COI, and LSU sequences from contemporary field-collected specimens revealed that H.munitum currently inhabits mesophotic rhodolith beds in the northwestern Gulf of Mexico, as well as the intertidal zone in the Florida Keys, Honduras, Atlantic Mexico, Caribbean Panama, and Guadeloupe, French West Indies. Species delimitation analyses reveal that the Western Atlantic and Australian H.munitum populations may be separate species. Two new species of Harveylithon from the northwestern Gulf of Mexico and one new species from the southwestern Gulf of Mexico, the Caribbean, and the Red Sea were also identified in the analyses and are described.

Numerical phenetic and phylogenetic relationships in silico among brown seaweeds (Phaeophyceae) from Gunungkidul, Yogyakarta, Indonesia

Abstract. Ningrum AM, Chasani AR. 2021. Numerical phenetic and phylogenetic relationships in silico among brown seaweeds (Phaeophyceae) from Gunungkidul, Yogyakarta, Indonesia. Biodiversitas 22: 3057-3064. Human activities such as industrial and tourism development on coastal areas in Indonesia are dreaded to affect seaweed diversity, including brown seaweeds (Phaeophyceae). Though study on brown seaweeds diversity has been done quite a lot, there is no record of analysis of phenetic and phylogenetic relationships among brown seaweeds. Hence, this study aims to determine the phenetic and phylogenetic relationships in silico among brown seaweeds and define characters that play a role in the clustering of brown seaweeds from Gunungkidul, Yogyakarta, Indonesia. Exploration was done using purposive sampling method. Numerical phenetic analysis was generated using MVSP 3.1. Further clustering method was implemented to identify phenetic relationships The PCA method was used to reveal morphological, anatomical, and biochemical characters that determine the clustering pattern. The phylogenetic relationships in silico analysis were conducted using rbcL genes from NCBI GenBank database. All multiple sequences were aligned using ClustalW and phylogram reconstruction was performed using Neighbor-Joining (NJ) method in MEGA 7.0. Our study showed that both the analyses, i.e., numerical phenetic and phylogenetic relationships in silico resulted in two main clusters although the species composition of the clusters was slightly different. The PCA analysis indicated that the morphological characters i.e. blade shape, phylloid shape, thalli height, length of the main axis, and the water bladder shape play an important role in the clustering of brown seaweeds species.

Watanabeales ord. nov. and twelve novel species of Trebouxiophyceae (Chlorophyta).

The Trebouxiophyceae include Chlorellales, Prasiolales, Trebouxiales, Microthamniales, and numerous members for which taxonomic locations have not been determined. The Watanabea clade is a group in the Trebouxiophyceae of Chlorella-like taxa, which are predominantly solitary and terrestrial. With the increase in descriptions of new species within the Watanabea clade in recent years, it is necessary to define the boundary and taxonomic status of the clade. In this study, nineteen strains of terrestrial algae collected from China were investigated based on molecular and morphological data, involving phylogenetic analyses, light and electron microscopy, and ITS-2 secondary structures. These 19 strains were described as five known species and 12 novel ones. Based on the consistent topological structures of the phylogenetic analyses for three taxonomic levels (the core Chorophyta, Trebouxiophyceae based on 18S sequences, Watanabeales and Symbiochloris based on 18S, 5.8S, ITS2 and rbcL gene sequences) and common morphological characteristics, especially the key phenotypic feature of unequally sized autospores, the Watanabea clade was defined as a new order, Watanabeales ord. nov., with one family, Watanabeaceae fam. nov. The boundary of Watanabeales was delineated and included the genera Chloroidium, Calidiella, Jaagichlorella, Kalinella, Massjukichlorella, Mysteriochloris, Polulichloris, Phyllosiphon, Watanabea, and Viridiella. Symbiochloris did not belong to Watanabeales according to the phylogenetic analyses and the reproduction by equal-sized autospores, aplanospores, or zoospores. The Watanabeales were separated into four groups during the phylogenetic analyses, but no regular differences in habitats or morphology could be used as the morphological basis to split Watanabeales into the four groups.

English

Apocynaceae sensu lato (s.l.) is treated as distinct family in some taxonomic systems while in others is treated as two separated families viz. Apocynaceae sensu stricto (s.s.) and Asclepiadaceae. So the main objective of the present study was to adopt one of the two views. The morphological characters viz. whole plant, lamina vein architecture and lamina epidermal characters of 20 wild and ornamental species were examined using LM and SEM. The data were numerically analyzed to detect the phenetic relationship among the studied species. DNA barcoding based on the sequencing of rbcL gene was cladistically analyzed to detect the phylogenetic relationship among the studied species. The generated phenogram showed a clear separation of two subseries; one of them belonged to Apocynaceae and the other with the remaining taxa belonging to Asclepiadaceae. The obtained cladogram showed that all of the studied species were divided into four lineages. It is concluded that the phenetic analysis supports the treatment of Apocynaceae s.l. as two distinct families; Apocynaceae s.s. Asclepiadaceae contrary of the phylogenetic one that supports the treatment of Apocynaceae s.s. and Asclepiadaceae as one family (Apocynaceae s.l.) with four subfamilies viz. Apocynoideae, Rauvolfiodeae, Asclepiadoideae and Periplocoideae. © 2021 Friends Science Publishers

On the nomenclatural reinstatement and lectotypification of Spyridia americana Durant (1850)

Abstract Southern New England and New York specimens of Spyridia ‘filamentosa’ were sequenced for the mitochondrial COI-5P and chloroplastic rbcL genes and determined to be distinct from Mediterranean (type locality) specimens of the same taxon. A little-known species name, Spyridia americana Durant, is applied to specimens collected from the northeastern coast of the United States.

New lineages of photobionts in Bolivian lichens expand our knowledge on habitat preferences and distribution of Asterochloris algae

We studied the biodiversity of Asterochloris photobionts found in Bolivian lichens to better understand their global spatial distribution and adaptation strategies in the context of a worldwide phylogeny of the genus. Based on nuclear ITS rDNA, the chloroplast rbcL gene and the actin type I gene we reconstructed a phylogenetic tree that recovered nine new Asterochloris lineages, while 32 Bolivian photobiont samples were assigned to 12 previously recognized Asterochloris lineages. We also show that some previously discovered Asterochloris photobiont species and lineages may occur in a broader spectrum of climatic conditions, and mycobiont species and photobionts may show different preferences along an altitude gradient. To reveal general patterns of of mycobiont specificity towards the photobiont in Asterochloris, we tested the influence of climate, altitude, geographical distance and effects of symbiotic partner (mycobiont) at the species level of three genera of lichen forming fungi: Stereocaulon, Cladonia and Lepraria. Further, we compared the specificity of mycobionts towards Asterochloris photobionts in cosmopolitan, Neotropical, and Pantropical lichen forming fungi. Interestingly, cosmopolitan species showed the lowest specificity to their photobionts, but also the lowest haplotype diversity. Neotropical and Paleotropical mycobionts, however, were more specific.

In Silico Analysis of rbcl Protein and D-Ribulose 1,5-bisphosphate Bond

The aim of this study was to determine the bond between the protein rbcL and D-Ribulose 1,5-bisphosphate by insilico. DNA sequences of the Mangrove Rhizophora mucronate rbcL gene DNA sequences from 7 different sources were obtained from NCBI for further alignment analysis using BioEdit software, phylogeny analysis using Mega6 software, molecular docking using PyRx software, preparation and visualization of docking results using Biovia Discovery Studio Visualizer software and analysis of the protein model quality based on the number of amino acid residues (Ramachandran plot analysis). The results of the docking molecular analysis showed interaction of 9 hydrogen bonds namely Asp203, Thr173, His294, Glu204, His327, Ser379, His298, Arg295, and Asn123 and 2 unfavorable bonds namely Lys177 and Lys175. This ligand and protein interaction complex was of good quality because the presence of amino acid residues in the most favored regions was greater than the amino acid residues in the disallowed regions outcomes

Phylogenetics of native conifer species in Vietnam based on two chloroplast gene regions rbcL and matK

We used two chloroplast gene regions (matK and rbcL) as a tool for the identification of 33 local conifer species. All 136 sequences, 101 newly generated (14 species for gene matK; 16 species for gene rbcL) and 35 retrieved from the GenBank, were used in the analysis. The highest genetic distance (matK region) was recorded between the species in Cupressaceae with an average of 5% (0.1–8.5), Podocarpaceae with an average of 6% (0–8.5), Taxaceae with an average of 5% (0.2–0.5) and Pinaceae with an average of 20.4% (0.8–54.1). The rbcL region showed a low genetic distance between the species in Cupressaceae 2% (0–3.3), Podocarpaceae 3% (0.6–3.4), Taxaceae 1% (0–2.1) and Pinaceae 1.2% (0–5.82). The phylogenetic analyses using the Maximum likelihood (ML) and Bayesian inference (BI) bootstrap values obtained at the branching nodes of each species ranged from 62 to 100% (Maximum likelihood bootstrap – MLBS and Bayesian posterior probabilities – BPP) for the matK gene; from 66 to 100% (MLBS) and 60 to 100% (BPP) for the rbcL region. The rbcL region was not identified between the species of Taxaceae and Cephalotaxaceae. The matK gene region was very clear in the different species among the families (Cupressaceae, Podocarpaceae, and Cephalotaxaceae) and unsuitable for identifying closely related species in Amentotaxus (Taxaceae) and Pinus (Pinaceae). The gene (matK) is a useful tool as a barcode in the identification of conifer species of Cupressaceae, Podocarpaceae, and Cephalotaxaceae in Vietnam.

Identification of three seagrass species in coral reef ecosystem by using multiple genes of DNA barcoding.

Seagrasses constitute a significant part of coral reef ecosystems, representing high primary productivity and one of the most important coastal habitats in marine ecosystems. Though seagrasses possess irreplaceable ecological services to the marine environment, taxonomical ambiguity still exists due to similar morphological characters and phenotypic plasticity. As an emerging technology, DNA barcoding can effectively identify cryptic species using a short orthologous DNA region. In this study, we collected samples from five different locations (Daya Bay, Xincun Bay, Sanya Bay, Xisha Islands, and Nansha Islands), and three seagrass species Cymodocea rotundata, Thalassia hemprichii and Halophila ovalis was evaluated. Moreover, ITS, matK and rbcL genes were used as DNA barcodes. The results indicated that single ITS and concatenated ITS/matK/rbcL both conducted better species resolution than single matK and rbcL. Nevertheless, single ITS was more convenient. Furthermore, in all the four topology trees, three species resolved as 3 clusters as well H. ovalis and T. hemprichii grouped as sister clade. In the meantime, differentiation lay in intra-species based on the result of single ITS and three-locus analysis. Within H. ovalis and T. hemprichii separately, individuals from Xisha Islands first group together, then grouped with individuals from Nansha Islands and/or Xincun Bay and/or Sanya Bay and/or Daya Bay, which indicated that geographical distribution influenced population evolution. However, intra-species differentiation did not emerge in the tree of matK or rbcL.

The First Report of Orobanche laxissima Parasitizing Pomegranate (Punica granatum) in Georgia.

Orobanche laxissima Uhlich & Rätzel (Orobanchaceae) is a polyphagous root parasitic plant distributed in the Caucasus Mountains and Transcaucasia; especially Russia, Georgia, Armenia, Azerbaijan, N.E. Turkey (Piwowarczyk et al. 2019). It infects many wild or sometimes cultivated trees and shrubs, such as Betulaceae, Oleaceae, Fagaceae, Aceraceae, Cornaceae, usually Fraxinus L., Fagus L., Carpinus L. (Piwowarczyk et al. 2019, 2020). Punica granatum L. (Lythraceae), commonly known as pomegranate, is native to the Caucasus, the Himalayas in North Pakistan and Northern India, and is widely cultivated, e.g. in USA and throughout the Mediterranean. Pomegranate is one of the first domesticated fruits and have been used in folk medicine or as a food for centuries. Fruit, seed, leaves, flower, root, or barks extracts have extensive medicinal properties (Shaygannia et al. 2015). Field surveys conducted in south-eastern Georgia in May 2019 revealed extensive infestations of O. laxissima on the roots of P. granatum in one locality in Kakheti Province, near Sighnaghi (41°37,4 N, 45°56,3 E, 480 m elevation), in roadside or hills scrub and cultivated areas of pomegranate. The infection was confirmed by verifying the attachment of the Orobanche to the Punica root. The population of the parasite consisted of at least a ca. thousand shoots, sometimes in one clump was ca. 100 individuals. A single plant of pomegranate was parasitized by few to c.a. a hundred of broomrape plants, and 10 to 20% of the ca. 1 ha location was infested. The main botanical features of the O. laxissima are: i) stem simple, (10-)25-40(-100) cm high, with haustoria; ii) inflorescence usually long to short cylindrical or lax, usually many-flowered; iii) calyx-segments entire or bidentate, rarely with 4 teeth; iv) corolla (16-)22-24(-31) mm long, tubular-bell-shaped; purple, pink, rarely dirty yellow, light brown; v) stigma purple, orange, or yellow (Piwowarczyk et al. 2019, 2020). For molecular analysis, total genomic DNA was extracted from the sample and the plastid gene rbcL (rubisco large subunit) was sequenced and amplified as described in Piwowarczyk et al. (2015). The sequence (1231 bp) was deposited in GenBank (MN384886). BLAST search found that it was most similar to (Query Cover 100%, Per Ident. 100%) O. laxissima (KR260928). To the best of our knowledge, this is the first report of a O. laxissima parasitizing P. granatum. O. laxissima appearing in large numbers on singles pomegranate shrubs can weaken the plants, and reduce flowering and fruiting. In the Caucasus region, O. laxissima was observed in mesophilic forests and shrubs, but our report suggests the possibility of a potential spread to neighboring cultivated areas, especially fruit trees and shrubs. Until now, only one report of pathogenic plants was documented for P. granatum, included Phelipanche aegyptiaca (Pers.) Pomel and O. crenata Forssk. in Israel (Dor et al. 2014).

Corallinapetrales and Corallinapetraceae: A new order and family of coralline red algae including Corallinapetra gabrielii comb. nov.

The coralline algal genus Corallinapetra is currently monospecific and was established on the species Corallinapetra novaezelandiae, known from a single collection from north-eastern New Zealand. On the basis of multi-gene phylogenetic analyses, Corallinapetra has been resolved apart from all currently recognized families and orders within the Corallinophycidae. We analyzed DNA sequence data from the holotype of Lithothamnion gabrielii, which has been considered a heterotypic synonym of L.muelleri, and an unidentified sample collected from Stewart Island in New Zealand, using psbA, rbcL, and COI-5P genes. We also observed detailed morpho-anatomical characters with light and scanning electron microscopy. Our phylogenetic analyses showed that L.gabrielii and the sample from New Zealand belonged to the same clade as Corallinapetra, distinct from other families and orders in the Corallinophycidae. Members of this clade are distinguishable from other families and orders in the Corallinophycidae by possessing sporangia that are surrounded by remnant sterile filaments that are weakly calcified in mature multiporate sporangial conceptacles that produce zonately divided tetrasporangia. Therefore, we propose that Corallinapetra be placed in its own family, Corallinapetraceae and order, Corallinapetrales, and that L.gabrielii should be assigned to Corallinapetra, as C.gabrielii, to reflect their phylogenetic relationships. We also obtained a partial rbcL sequence data from the lectotype of L.muelleri, the generitype of Lithothamnion. Comparison of the L.muelleri type sequence with L.gabrielii unambiguously demonstrated that these two species are not conspecific, and confirm the placement of L.muelleri within the Hapalidiales.

Comparative and phylogenetic analysis of the complete chloroplast genome sequences of Lactuca raddeana and Lactuca sativa

Lactuca raddeana is a biennial plant of the Lactuca genus belonging to the Asteraceae family. The classification of Lactuca is controversial, and no consistent conclusions have been reached based on the analysis of morphological characters and different molecular markers. Here, we sequenced and assembled the complete chloroplast genome of L. raddeana. This genome has a total length of 152,339 bp, a conservative quartile structure that is composed of a large single-copy (LSC) region of 83,976 bp, a small-copy (SSC) region of 18,521 bp, and a pair of inverted repeats (IRs) of 24,921 bp. The genome contains 112 unique genes, including 79 protein-coding, four rRNA, and 29 tRNA genes. Repeat analysis obtained 17 microsatellite, 16 tandem, and 17 interspersed repeats. Comparison of sequence divergence between L. raddeana and L. sativa found the intergenic spacer trnC-petN exhibited the highest degree of variation. Three phylogenetic trees based on the 72 shared protein, matK gene, and rbcL gene sequences showed that L. raddeana and L. sativa are clustered together. The acquisition and comparative analysis of the chloroplast genome provide a valuable resource for the taxonomic and phylogenetic studies of Lactuca.

Production of the sheep pox virus structural protein SPPV117 in tobacco chloroplasts

ObjectiveA chloroplast transgenic approach was assessed in order to produce a structural protein SPPV117 of sheep pox virus in Nicotiana tabacum for the future development of a plant-based subunit vaccine against sheep pox.ResultsTwo DNA constructs containing SPPV117 coding sequence under the control of chloroplast promoter and terminator of psbA gene or rrn promoter and rbcL terminator were designed and inserted into the chloroplast genome by a biolistic method. The transgenic plants were selected via PCR analysis. Northern and Western blot analysis showed expression of the transgene at transcriptional and translational levels, respectively. The recombinant protein accumulated to about 0.3% and 0.9% of total soluble protein in leaves when expressed from psbA and rrn promoter, respectively. Plant-produced SPPV117 protein was purified using metal affinity chromatography and the protein yield was 19.67 ± 1.25 µg g−1 (FW). The serum of a sheep infected with the virus recognised the chloroplast-produced protein indicating that the protein retains its antigenic properties.ConclusionsThese results demonstrate that chloroplasts are a suitable system for the production of a candidate subunit vaccine against sheep pox.