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dna-barcoding of Some Medicinal Plant Species in Saudi Arabia Using rbcL and matK Genes

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Phyton-International Journal of Experimental Botany 2020年第4期89卷 1059-1081页

作者： Abdulrahman Alasmari Department of Biology Faculty of ScienceUniversity of TabukTabuk71491Saudi Arabia

In the Kingdom of Saudi Arabia(KSA),thousands of plants are considered to have therapeutic value.The ambiguous use of identification mainly morphological characteristics of many plants has resulted in the adulteration and displacement of plant products which undermine their therapeutic value and weak documentation of plant resources.The aims of this study were therefore to evaluate genetic variability and explore the phylogeographic architecture for Saudi medicinal plant samples using rbcL and matK genes as barcodes for genomic identification.The matK and rbcL sequences collected for these samples were used as key markers for examining the relationship between Saudi medicinal plant species based on genetic diversity.During our study we were successful in identifying and documenting 4 different species(Foeniculum vulgare,Nitraria retusa,Dodonaea viscosa,and Rumex nervosus)located in Saudi Arabia using dna barcoding technique.A total number of 8 sequences were obtained with a total sequence length of 6176 bp,where it ranged from 617 bp to 878 bp with an aver-age length of 772 bp.The total number of rbcL sequences length is 2801 bp,where it ranges from 617 bp to 807 bp with an average length of 700.2 bp.Out of the 4 plant samples used,only three samples were identified correctly on the species level with an identity percentage higher than 95%using rbcL gene.Additionally,4 matK sequences have been retrieved belong to 4 species.The total number of matK sequences length is 3375 bp,where it ranges from 819 bp to 878 bp with an average length of 843.8 bp.Out of the 4 plant samples used,only two samples were identified correctly on the species level with an identity percentage higher than 98% using matK gene.Both rbcL and matK have been able to identify most of our collected plant samples by genus,and some by species.Using only one dna-barcoding technique was not reliable for plant identification,where matK and rbcL must be used as a dual dna-barcoding procedure.

关键词： Phylogeographic architecture genomic identification dna barcoding

Morphology versus dna barcoding： two sides of the same coin. A case study of Ceutorhynchus erysimi and C. contractus identification

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Insect Science 2016年第4期23卷 638-648页

作者： Svetlana Stepanovic Andrea Kosovac Oliver Krstic Jelena Jovic lvo Tosevski Faculty of Science Institute of Biology and Ecology University of Kragujevac 34000 Kragujevac Serbia Department of Plant Pests Institute for Plant Protection and Environment Banatska 33 11080 Zemun Serbia CABI 1 Rue des Grillons 2800 Delemont Switzerland

Genotyping of 2 well-known weevil species from the genus Ceutorhynchus （Coleoptera： Curculionidae） distributed in west Palearctic, C. erysimi and C. contractus, revealed phenotype versus genotype inconsistencies in a set of 56 specimens （25 C. erysimi and 31 C. contractus） collected from 25 locations in Serbia and Montenegro. An analysis of the mitochondrial cytochrome oxidase subunit I gene （CO1）, widely used as a barcoding region, and a nuclear gene, elongation factor-v （EF-1α）, revealed stable genetic divergence among these species. The average uncorrected pairwise distances for the CO1 and EF-1α genes were 3.8%, and 1.3%, respectively, indicating 2 genetically well-segregated species. However, the genetic data were not congruent with the phenotypic characteristics of the studied specimens. In the first place, C. erysimi genotypes were attached to specimens with phenotypic characteristics of C. contractus. Species-specific PCR-RFLP assays for the barcoding gene COIwere applied for the molecular identification of 101 additional specimens of both morphospecies （33 C. erysimi and 68 C. contractus） and were found to confirm this incongruity. The discrepancy between the genetic and morphological data raises the question of the accuracy of using a barcoding approach, as it may result in misleading conclusions about the taxonomic position of the studied organism. Additionally, the typological species concept shows considerable weakness when genetic data are not supported with phenotypic characteristics as in case of asymmetric introgression, which may cause certain problems, especially in applied studies such as biological control programs in which the biological properties of the studied organisms are the main focus.

关键词： Ceutorhynchus contractus Ceutorhynchus erysimi dna barcoding molecular identification morphology PCR-RFLP

New universal matK primers for dna barcoding angiosperms

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Journal of Systematics and Evolution 2011年第3期49卷 176-181页

作者： 1,2Jing YU 1Jian-Hua XUE 1Shi-Liang ZHOU* 1(State Key Laboratory of Systematic and Evolutionary Botany,Institute of Botany,Chinese Academy of Sciences,Beijing 100093,China) 2(Graduate University of the Chinese Academy of Sciences,Beijing 100049,China) (State Key Laboratory of Systematic and Evolutionary Botany Institute of Botany Chinese Academy of Sciences Beijing 100093 China) (Graduate University of the Chinese Academy of Sciences Beijing 100049 China)

The chloroplast maturase K gene(matK) is one of the most variable coding genes of angiosperms and has been suggested to be a "barcode" for land plants.However,matK exhibits low amplification and sequencing rates due to low universality of currently available primers and mononucleotide repeats.To resolve these technical problems,we evaluated the entire matK region to find a region of 600-800 bp that is highly variable,represents the best of all matK regions with priming sites conservative enough to design universal primers,and avoids the mononucleotide repeats.After careful evaluation,a region in the middle was chosen and a pair of primers named matK472F and matK1248R was designed to amplify and sequence the matK fragment of approximately 776 bp.This region encompasses the most variable sites,represents the entire matK region best,and also exhibits high amplification rates and quality of sequences.The universality of this primer pair was tested using 58 species from 47 families of angiosperm plants.The primers showed a strong amplification(93.1%) and sequencing(92.6%) successes in the species tested.We propose that the new primers will solve,in part,the problems encountered when using matK and promote the adoption of matK as a dna barcode for angiosperms.

关键词： angiosperm dna barcoding matK primer.

Molecular Identification of Dried Shellfish Products Sold on the Market Using dna barcoding

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Journal of Ocean University of China 2021年第4期20卷 931-938页

作者： SUN Shao’e ZHANG Xiaojie KONG Lingfeng LI Qi Key Laboratory of Mariculture Ministry of EducationOcean University of ChinaQingdao 266003China Laboratory for Marine Fisheries Science and Food Production Processes Pilot National Laboratory for Marine Science and Technology(Qingdao)Qingdao 266237China

The dried shellfish products with rich nutrients and low-calorie content are favorite food in China,especially in coastal areas.However,the species of dried shellfish products in the market are usually unknown,as the taxonomic features were removed during the production process.This study described the application of dna barcoding technique to the identification of 100 dried shellfish(scallop,squid,octopus and cuttlefish)products in markets.Samples were authenticated by comparing mitochondrial cytochrome oxidase subunit I(COI)gene and 16S ribosomal RNA(16S rRNA)gene sequences with public reference taxonomic databases.The results showed that all the 100 products can be identified at species level.Sixty four scallop adductor products were processed using the bay scallop,Argopecten irradians,and one was from Portuguese oyster,Crassostrea angulata.All the 27 squid,2 cuttlefish and 6 octopus products were produced by the Jumbo flying squid,Dosidicus gigas.The neighbour-joining tree is in agreement with the results of dna barcoding analysis.The 64 scallop samples formed one A.irradians cluster,leaving Sca65 clustered with the reference oyster sequence C.angulata(MH997922).All the 35 cephalopod(squid,octopus and cuttlefish)samples formed a D.gigas cluster.This investigation revealed a low variety of dried shellfish products sold on the market,and highlighted the high rate of mislabeling and species substitution.Our present work provides a practical method for tracing and authenticating shellfish products.

关键词： dried shellfish product dna barcoding species identification mislabeling species substitution

Identification of species within Tetrastigma (Miq.) Planch.(Vitaceae) based on dna barcoding techniques

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Journal of Systematics and Evolution 2011年第3期49卷 237-245页

作者： 1,2Yuan-Miao FU# 1,2Wei-Mei JIANG# 1,2Cheng-Xin FU* 1(The Key Laboratory of Conservation Biology for Endangered Wildlife of the Ministry of Education,College of Life Sciences,Zhejiang University,Hangzhou 310058,China) 2(Laboratory of Systematic and Evolutionary Botany and Biodiversity,Institute of Plant Sciences and Conservation Center for Gene Resources of Endangered Wildlife,Zhejiang University,Hangzhou 310058,China) (The Key Laboratory of Conservation Biology for Endangered Wildlife of the Ministry of Education College of Life Sciences Zhejiang University Hangzhou 310058 China) (Laboratory of Systematic and Evolutionary Botany and Biodiversity Institute of Plant Sciences and Conservation Center for Gene Resources of Endangered Wildlife Zhejiang University Hangzhou 310058 China)

Many species of Tetrastigma(Miq.) Planch.(Vitaceae) have long been used as medicinal plants in China,and some are endangered due to overexploitation.Although adulterants are often added to traditional Chinese medicines,there is no reliable or practical method for identifying them.In this study,we used four markers(rbcL,matK,trnH-psbA,and internal transcribed spacer [ITS]) as dna barcodes to test their ability to distinguish species of Tetrastigma.The results indicated that the best barcode was ITS,which showed significant inter-specific genetic variability,and thus its potential as a dna barcode for identifying Tetrastigma.Multiple loci provided a greater ability to distinguish species than single loci.We recommend using the combined rbcL+matK+ITS barcode for the genus.Phylogenetic trees from each barcode were compared.Analyses using the unweighted pair group method with arithmetic mean discriminated an equal or greater percentage of resolvable species than did neighbor joining,maximum likelihood,or maximum parsimony analyses.Additionally,five medicinal species of Tetrastigma,especially T.hemsleyanum,could be identified precisely using dna barcoding.

关键词： dna barcoding medicinal plants species identification Tetrastigma.

Phylogenetic Analysis of Brine Shrimp (Artemia) in China Using dna barcoding

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Genomics, Proteomics & Bioinformatics 2008年第3期6卷 155-162页

作者： Weiwei Wang Qibin Luo Haiyan Guo Peter Bossier Gilbert Van Stappen Patrick Sorgeloos Naihong Xin Qishi Sun Songnian Hu Jun Yu Key Laboratory of Genome Sciences and Information Beijing Institute of Genomics Chinese Academy of Sciences Beijing 100029 China Graduate University of Chinese Academy of Sciences Beijing 100049 China James D. Watson Institute of Genome Sciences Zhejiang University Hangzhou 310008 China School of Traditional Chinese Materia Medica Shenyang Pharmaceutical University Shenyang 110016 China Laboratory of Aquaculture ＆ Artemia Reference Center Ghent University B-9000 Ghent Belgium Salt Research Institute Tianjin 300450 China.

dna barcoding is a powerful approach for characterizing species of organisms, especially those with almost identical morphological features, thereby helping to to establish phylogenetic relationships and reveal evolutionary histories. In this study, we chose a 648-bp segment of the mitochondrial gene, cytochrome c oxidase subunit 1 （COI）, as a standard barcode region to establish phylogenetic relationships among brine shrimp （Artemia） species from major habitats around the world and further focused on the biodiversity of Artemia species in China, especially in the Tibetan Plateau. Samples from five major salt lakes of the Tibetan Plateau located at altitudes over 4,000 m showed clear differences from other Artemia populations in China. We also observed two consistent amino acid changes, 153A/V and 183L/F, in the COI gene between the high and low altitude species in China. Moreover, indels in the COI sequence were identified in cyst and adult samples unique to the Co Qen population from the Tibetan Plateau, demonstrating the need for additional investigations of the mitochondrial genome among Tibetan Artemia populations.

关键词： Artemia dna barcoding molecular phylogeny Tibetan Plateau

Species delimitation in the green algal genus Codium (Bryopsidales) from Korea using dna barcoding

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Acta Oceanologica Sinica 2015年第4期34卷 114-124页

作者： LEE Hyung Woo KIM Myung Sook Department of Biology Jeju National University

Codium, one of the largest marine green algal genera, is difficult to delimit species boundary accurately based on morphological identification only. dna barcoding is a powerful tool for discriminating species of seaweeds. The plastid elongation factor TU （tufA） is considered as maker to perform dna barcoding of green algal species than rbcL gene due to universality and rapid evolution rate. We conducted dna barcoding application to Codium specimens from the Jeju Island, Korea to overcome the limit of morphological identification and to confirm the species diversity. As a result of applying tufA marker, we newly generated fifty-five tufA barcodes to resolve eight species. TufA marker exhibited 6.1%–21.8% interspecific divergences, wider than the gap of rbcL exon 1, 3.5%–11.5%. Molecular analysis of rbcL exon 1 sequences of Codium revealed eight distinct species like tufA analysis separated in five phylogenetic groups. dna barcoding of the genus Codium using tufA marker is more helpful to overcome the limit of morphological identification, and this is more potential to reveal cryptic species and to resolve the relationships among subspecies than rbcL analysis alone. The complement of tufA barcoding and rbcL analyses including morphology for the genus Codium in the northwestern Pacific will give much more reliable achievement for discovering species diversity and resolving the phylogenetic relationships.

关键词： Codium dna barcoding rbcL Species delimitation tufA

16S rRNA is a better choice than COI for dna barcoding hydrozoans in the coastal waters of China

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Acta Oceanologica Sinica 2014年第4期33卷 55-76页

作者： ZHENG Lianming HE Jinru LIN Yuanshao CAO Wenqing ZHANG Wenjing College of Ocean and Earth Sciences Xiamen University Marine Biodiversity and Global Change Research Center Xiamen University Fujian Provincial Key Laboratory for Coastal Ecology and Environmental Studies (CEES) XiamenUniversity

Identification of hydrozoan species is challenging, even for taxonomic experts, due to the scarcity of distinct morphological characters and phenotypic plasticity. dna barcoding provides an efficient method for species identification, however, the choice between mitochondrial cytochrome c oxidase subunit I（COI） and large subunit ribosomal RNA gene（16S） as a standard barcode for hydrozoans is subject to debate. Herein, we directly compared the barcode potential of COI and 16S in hydrozoans using 339 sequences from 47 pelagic hydrozoan species. Analysis of Kimura 2-parameter genetic distances（K2P） documented the mean intraspecific/interspecific variation for COI and 16S to be 0.004/0.204 and 0.003/0.223, respectively. An obvious ＂barcoding gap＂ was detected for all species in both markers and all individuals of a species clustered together in both the COI and 16S trees. These results suggested that the species within the studied taxa can be efficiently and accurately identified by COI and 16S. Furthermore, our results confirmed that 16S was a better phylogenetic marker for hydrozoans at the genus level, and in some cases at the family level. Considering the resolution and effectiveness for barcoding and phylogenetic analyses of Hydrozoa, we strongly recommend 16S as the standard barcode for hydrozoans.

关键词： dna barcoding hydrozoan COI 16S rRNA

Identification of Kalidium species(Chenopodiaceae)by dna barcoding

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Research in Cold and Arid Regions 2017年第1期9卷 89-96页

作者： XiaoHui Liang YuXia Wu State Key Laboratory of Grassland Agro-Ecosystem School of Life SciencesLanzhou UniversityLanzhouGansu 730000China

dna barcoding is an increasingly prevalent molecular biological technology which uses a short and conserved dna fragment to facilitate rapid and accurate species identification. Kalidium species are distributed in saline soil habitat throughout Southeast Europe and Northwest Asia, and used mainly as forage grass in China. The discrimination of Ka-lidium species was based only on morphology-based identification systems and limited to recognized species. Here, we tested four dna candidate loci, one nuclear locus （ITS, internal transcribed spacer） and three plastid loci （rbcL9 matK and ycflb）, to select potential dna barcodes for identifying different Kalidium species. Results showed that the best dna barcode was ITS locus, which displayed the highest species discrimination rate （100%）, followed by matK （33.3%）,ycflb （16.7%）, and rbcL （16.7%）. Meanwhile, four loci clearly identified the variant species, Kalidium cuspidatum （Ung.-Stemb.） Gmb.var.A. J. Li,as a single species in Kalidium.

关键词： dna barcoding Kalidium species identification