Authentication of Herbal Medicines Dipsacus asper and Phlomoides umbrosa Using DNA Barcodes, Chloroplast Genome, and Sequence Characterized Amplified Region (SCAR) Marker

Dried roots of Dipsacus asper (Caprifoliaceae) are used as important traditional herbal medicines in Korea. However, the roots are often used as a mixture or contaminated with Dipsacus japonicus in Korean herbal markets. Furthermore, the dried roots of Phlomoides umbrosa (Lamiaceae) are used indiscriminately with those of D. asper, with the confusing Korean names of Sok-Dan and Han-Sok-Dan for D. asper and P. umbrosa, respectively. Although D. asper and P. umbrosa are important herbal medicines, the molecular marker and genomic information available for these species are limited. In this study, we analysed DNA barcodes to distinguish among D. asper, D. japonicus, and P. umbrosa and sequenced the chloroplast (CP) genomes of D. asper and D. japonicus. The CP genomes of D. asper and D. japonicus were 160,530 and 160,371 bp in length, respectively, and were highly divergent from those of the other Caprifoliaceae species. Phylogenetic analysis revealed a monophyletic group within Caprifoliaceae. We also developed a novel sequence characterised amplified region (SCAR) markers to distinguish among D. asper, D. japonicus, and P. umbrosa. Our results provide important taxonomic, phylogenetic, and evolutionary information on the Dipsacus species. The SCAR markers developed here will be useful for the authentication of herbal medicines.

Download Full-text

Development of SCAR Markers Based on Improved RAPD Amplification Fragments and Molecular Cloning for Authentication of Herbal Medicines Angelica sinensis, Angelica acutiloba and Levisticum officinale

Natural Product Communications ◽

10.1177/1934578x1501001027 ◽

2015 ◽

Vol 10 (10) ◽

pp. 1934578X1501001 ◽

Cited By ~ 1

Author(s):

Chun Zhang ◽

Zhiqiang Mei ◽

Jingliang Cheng ◽

Yin He ◽

Md. Asaduzzaman Khan ◽

...

Keyword(s):

Molecular Cloning ◽

Scar Marker ◽

Herbal Medicines ◽

Pcr Amplification ◽

Angelica Sinensis ◽

Scar Markers ◽

Helpful Tool ◽

Sequence Characterized Amplified Region ◽

Ecological Conservation ◽

Angelica Acutiloba

Molecular cloning from DNA fragments of improved RAPD amplification of Angelica sinensis, Angelica acutiloba and Levisticum officinale, provided novel sequence-characterized amplified region (SCAR) markers A13, A23, Al-34 and Al-0 whose sequences were deposited in the GenBank database with the accession numbers KP641315, KP641316, KP641317 and KP641318, respectively. By optional PCR amplification, the SCAR markers A13 and A23 are Levisticum officinale-specific, whereas the SCAR marker Al-34 is Angelica acutiloba-specific, and the SCAR marker Al-0 is Angelica sinensis-specific. These diagnostic SCAR markers may be useful for genetic authentications, for ecological conservation of all three medicinal plants and as a helpful tool for the genetic authentication of adulterant samples.

Download Full-text

PCR-based molecular markers for identification of taxa from the Calypogeia fissa complex (Jungermanniopsida, Calypogeiaceae)

Biodiversity Research and Conservation ◽

10.2478/v10119-012-0024-3 ◽

2012 ◽

Vol 28 (1) ◽

pp. 9-18 ◽

Cited By ~ 1

Author(s):

Katarzyna Buczkowska ◽

Patrycja Gonera ◽

Bartosz Hornik

Keyword(s):

Molecular Markers ◽

Dna Sequences ◽

Scar Marker ◽

Issr Markers ◽

Genetic Differences ◽

Scar Markers ◽

Sequence Characterized Amplified Region ◽

Pcr Product ◽

Isozyme Loci ◽

Diagnostic Traits

Abstract Within Calypogeia fissa, two subspecies connected with geographic distribution are formally recognized: C. fissa subsp.fissa in Europe and C. fissa subsp.neogea in North America. Isoenzyme studies have shown that the European subspecies is genetically differentiated and composed of three genetically distinct groups PS, PB and G. The PS group has the most distinctive morphological features, but no morphological diagnostic traits have been found for groups PB and G. The sequence characterized amplified region (SCAR) markers developed on the basis of ISSR markers, applied in the study, allowed the delimitation of all groups distinguished in Europe within the C. fissa complex (PS, PB and G). The markers also revealed genetic differences between the European and American subspecies. Five primer pairs (Cal01, Cal03-Cal06) of the six pairs studied are useful as the diagnostic tool for the identification of particular groups from the C.fissa complex. The examined SCAR markers showed that the PS group of C.fissa subsp.fissa was the most distinct; it differed from both groups PB and G as well as from C.fissa subsp.neogea. All plants determined on the basis of diagnostic isozyme loci as the PS group amplified a longer product (380 bp) of the Cal04 primer pair than the rest of studied groups and yielded no amplification products in Cal03, Cal05 and Cal06 primers. The primer pair Cal03 distinguished the plants of the PB group from the remaining groups, since only the PB group generated a PCR product of about 290 bp. The genetic differences between all four studied groups of the C.fissa complex were supported by DNA sequences of the SCAR marker Cal04.

Download Full-text

Efficiency of Randomly Amplified Polymorphic DNA to Sequence Characterized Amplified Region Marker Conversion and their Comparative Polymerase Chain Reaction Sensitivity in Cucumber

Journal of the American Society for Horticultural Science ◽

10.21273/jashs.124.2.128 ◽

1999 ◽

Vol 124 (2) ◽

pp. 128-135 ◽

Cited By ~ 30

Author(s):

Thomas Horejsi ◽

Jodie M. Box ◽

Jack E. Staub

Keyword(s):

Rapd Markers ◽

Scar Marker ◽

Scar Markers ◽

Sequence Characterized Amplified Region ◽

Pcr Product ◽

Rapd Pcr ◽

Pcr Products ◽

Polymerase Chain ◽

Template Dna ◽

Pcr Conditions

The conversion of randomly amplified polymorphic DNA (RAPD) markers to sequence characterized amplified region (SCAR) markers, and the effects of differing polymerase chain reaction (PCR) conditions were studied in cucumber (Cucumis sativus L.). Attempts were made to clone and sequence 75 RAPD PCR products to produce SCAR primers (16 to 22 nucleotides) designed to amplify original RAPD PCR products. The influence of template DNA source, purity, and concentration, MgCl2 concentration, Taq polymerase source, and type of thermocycler upon RAPD and SCAR marker performance was evaluated. Conversion of RAPD to SCAR markers was not universally successful, and SCAR primers reacted differently to varying PCR conditions. Only 48 (64%) of 75 RAPD markers were successfully converted to SCAR markers and 11 (15%) of these reproduced the polymorphism observed with the original RAPD PCR product. Moreover, some SCAR primer pairs produced multiple polymorphic PCR products. The band intensity of SCAR markers were brighter (P = 0.05) than their corresponding RAPD markers with only one exception. The SCAR markers examined were less influenced (P = 0.05) by MgCl2 concentration than their corresponding RAPD markers. However, some SCAR markers were more sensitive to reaction impurities than their RAPD counterparts and SCAR markers tended to be less readily visualized (decrease in frequency of visible PCR product) with low concentrations (1 and 2 mm) of template DNA than their corresponding RAPD markers. Neither the source of Taq nor the type of thermocycler used affected the performance of SCAR and RAPD markers. These data suggest that although SCAR markers may demonstrate enhanced performance over the RAPD markers from which they are derived, careful consideration must be given to both the costs and potential benefits of SCAR marker development in cucumber.

Download Full-text

Identification of the B, Q, and native Brazilian biotypes of the Bemisia tabaci species complex using Scar markers

Pesquisa Agropecuária Brasileira ◽

10.1590/s0100-204x2016000500016 ◽

2016 ◽

Vol 51 (5) ◽

pp. 555-562 ◽

Cited By ~ 3

Author(s):

Paulo Roberto Queiroz ◽

Erica Soares Martins ◽

Nazaré Klautau ◽

Luzia Lima ◽

Lilian Praça ◽

...

Keyword(s):

Bemisia Tabaci ◽

Species Complex ◽

Scar Marker ◽

Random Amplified Polymorphic Dna ◽

Scar Markers ◽

Sequence Characterized Amplified Region ◽

Field Samples ◽

Scar Primer ◽

Q Biotype ◽

B Biotype

Abstract: The objective of this work was to develop sequence-characterized amplified region (Scar) markers to identify the B, Q, and native Brazilian biotypes of the sweet potato whitefly [Bemisia tabaci (Hemiptera: Aleyrodidae)]. Random amplified polymorphic DNA (RAPD) amplification products, exclusive to the B and Brazilian biotypes, were selected after the analysis of 12,000 samples, in order to design a specific Scar primer set. The BT-B1 and BT-B3 Scar markers, used to detect the B biotype, produced PCR fragments of 850 and 582 bp, respectively. The BT-BR1 Scar marker, used to identify the Brazilian biotype, produced a PCR fragment of 700 bp. The Scar markers were tested against the Q biotype, and a flowchart was proposed to indicate the decision steps to use these primers, in order to correctly discriminate the biotypes. This procedure allowed to identify the biotypes that occur in field samples, such as the B biotype. The used set of primers allowed to discriminate the B, Q, and native Brazilian biotypes of B. tabaci. These primers can be successfully used to identify the B biotype of B. tabaci from field samples, showing only one specific biotype present in all cultures.

Download Full-text

Development of SCAR markers for sex determination in the dioecious shrub Aucuba japonica (Cornaceae)

Genome ◽

10.1139/g08-120 ◽

2009 ◽

Vol 52 (3) ◽

pp. 231-237 ◽

Cited By ~ 12

Author(s):

Masayuki Maki

Keyword(s):

Sex Identification ◽

The Other ◽

Scar Markers ◽

Specific Sequence ◽

Sequence Characterized Amplified Region ◽

Annealing Temperatures ◽

Aucuba Japonica ◽

Males And Females ◽

Male Specific ◽

Scar Primers

Two sex-linked fragments were identified by RAPD analyses in the dioecious diploid shrub Aucuba japonica var. ovoidea and were converted into markers of male-specific sequence characterized amplified region (SCAR) markers. PCRs using the primers designed in this study correctly discriminated 24 flowering males and 24 flowering females at higher annealing temperatures (SCAR markers OPA10-424 at 55 °C and OPN11-1095 at 65 °C), although at relatively low annealing temperatures, the fragments were amplified in both males and females. These SCAR primers were also tested to see whether they were applicable to sex identification in the conspecific tetraploid Aucuba japonica var. japonica. One set pf SCAR primers could be used for sex identification even in this tetraploid variety, although the other failed. The SCAR markers developed in this study will provide a powerful tool in identifying the sex of immature plants of dioecious A. japonica, which is a commercially valuable shrub due to its conspicuous fruits.

Download Full-text

Genetic diversity of root-knot nematodes from Brazil and development of SCAR markers specific for the coffee-damaging species

Genome ◽

10.1139/g02-054 ◽

2002 ◽

Vol 45 (5) ◽

pp. 862-870 ◽

Cited By ~ 90

Author(s):

Onivaldo Randig ◽

Michel Bongiovanni ◽

Regina M.D.G Carneiro ◽

Philippe Castagnone-Sereno

Keyword(s):

Genetic Diversity ◽

Multiplex Pcr ◽

Rapd Markers ◽

Phylogenetic Analyses ◽

The Other ◽

Scar Markers ◽

Meloidogyne Hapla ◽

Root Knot Nematodes ◽

Sequence Characterized Amplified Region ◽

Scar Primers

RAPD markers were used to characterize the genetic diversity and relationships of root-knot nematodes (RKN) (Meloidogyne spp.) in Brazil. A high level of infraspecific polymorphism was detected in Meloidogyne arenaria, Meloidogyne exigua, and Meloidogyne hapla compared with the other species tested. Phylogenetic analyses showed that M. hapla and M. exigua are more closely related to one another than they are to the other species, and illustrated the early divergence of these meiotically reproducing species from the mitotic ones. To develop a PCR-based assay to specifically identify RKN associated with coffee, three RAPD markers were further transformed into sequence-characterized amplified region (SCAR) markers specific for M. exigua, Meloigogyne incognita and Meloidogyne paranaensis, respectively. After PCR using the SCAR primers, the initial polymorphism was retained as the presence or absence of amplification. Moreover, multiplex PCR using the three pairs of SCAR primers in a single reaction enabled the unambiguous identification of each species, even in mixtures. Therefore, it is concluded that the method developed here has potential for application in routine diagnostic procedures.Key words: diagnostic, multiplex PCR, phylogeny, RAPD, root-knot nematodes.

Download Full-text

Development of RAPD-SCAR markers for Lonicera japonica Thunb. (Caprifolicaceae) variety authentication by improved RAPD and DNA cloning

Revista de Biología Tropical ◽

10.15517/rbt.v62i4.13493 ◽

2014 ◽

Vol 62 (4) ◽

pp. 1649 ◽

Cited By ~ 13

Author(s):

Luquan Yang ◽

Md. Asaduzzaman Khan ◽

Zhiqiang Mei ◽

Manman Yang ◽

Tiandan Zhang ◽

...

Keyword(s):

Molecular Markers ◽

Molecular Marker ◽

Scar Marker ◽

Rapd Analysis ◽

Random Amplified Polymorphic Dna ◽

Vital Role ◽

Lonicera Japonica ◽

Scar Markers ◽

Sequence Characterized Amplified Region ◽

Young Leaves

Genetic diversity within a species is a common feature, which plays a vital role in its survival and adaptability, and is important for the identification and authentication of a species. Lonicera japonica is a traditionally used medicinal plant, which have been recently genetically characterized by an improved random ampliﬁed polymorphic DNA (RAPD) analysis. In this study, the molecular markers on the basis of these RAPD fragments have been developed to identify specific L. japonica variety. The DNAs were extracted from fresh young leaves of different samples of L. japonica collected from Shenzhen, Yichang, Leshan, Emei and Loudi, China. The DNA materials were amplified using improved RAPD PCR. Different RAPD bands were excised, cloned and developed for stable sequence-characterized amplified region (SCAR) markers with different species. Two SCAR markers, JYH3-3 and JYH4-3, have been successfully cloned from improved RAPD fragments. The SCAR marker JYH3-3 was found specific for all of the L. japonica samples collected from the different regions, and another marker JYH 4-3 was strictly specific to the Shenzhen sample from Guangdong province, which is geographically distant from Hubei, Sichuan and Hunan Provinces (source of other L. japonica samples). The marker JYH3-3 was found as specific molecular marker for the identification of L. japonica, while JYH4-3 was found as molecular marker strictly specific for the Shenzhen sample. The developed SCAR markers might serve as more specific molecular markers for L. japonica variety authentication. The combination of improved RAPD analysis and SCAR marker development have resulted useful tools to study the genetic variety of any organism, which we have successfully applied here in L. japonica.de cualquier organismo, que hemos aplicado con éxito en L. japonica.

Download Full-text

Development of Randomly Amplified Polymorphic DNA Based SCAR Marker for Identification ofIpomoea mauritianaJacq (Convolvulaceae)

Evidence-based Complementary and Alternative Medicine ◽

10.1093/ecam/neq023 ◽

2011 ◽

Vol 2011 ◽

pp. 1-6 ◽

Cited By ~ 7

Author(s):

Kambiranda Devaiah ◽

Subramani Paranthaman Balasubramani ◽

Padma Venkatasubramanian

Keyword(s):

Scar Marker ◽

The Other ◽

Herbal Drug ◽

Randomly Amplified Polymorphic Dna ◽

Pueraria Tuberosa ◽

Sequence Characterized Amplified Region ◽

Cycas Circinalis ◽

Scar Primers

Vidari is an Ayurvedic herbal drug used as aphrodisiac, galactagogue and is also used in the preparation ofChyavanaprash. Tubers ofIpomoea mauritianaJacq. (Convolvulaceae),Pueraria tuberosa(Roxb. ex Willd.) DC (Fabaceae),Adenia hondala(Gaertn.) de Wilde (Passifloraceae) and pith ofCycas circinalisL. (Cycadaceae) are all traded in the name of Vidari, creating issues of botanical authenticity of the Ayurvedic raw drug. DNA-based markers have been developed to distinguishI. mauritianafrom the other Vidari candidates. A putative 600-bp polymorphic sequence, specific toI. mauritianawas identified using randomly amplified polymorphic DNA (RAPD) technique. Furthermore, sequence characterized amplified region (SCAR) primers (IM1F and IM1R) were designed from the unique RAPD amplicon. The SCAR primers produced a specific 323-bp amplicon in authenticI. mauritianaand not in the allied species.

Download Full-text

Clinical Use of Toxic Proteins and Peptides from Tian Hua Fen and Scorpion Venom

Current Protein and Peptide Science ◽

10.2174/1389203719666180622100641 ◽

2019 ◽

Vol 20 (3) ◽

pp. 285-295

Author(s):

Chen Ling ◽

Yuanhui Zhang ◽

Jun Li ◽

Wenli Chen ◽

Changquan Ling

Keyword(s):

Herbal Medicines ◽

Scorpion Venom ◽

The Other ◽

Western World ◽

Chinese Government ◽

Clinical Use ◽

Potential Benefits ◽

Scientific Results ◽

Proteins And Peptides ◽

Toxic Proteins

Traditional Chinese Medicine (TCM) has been practiced in China for thousands of years. As a complementary and alternative treatment, herbal medicines that are frequently used in the TCM are the most accepted in the Western world. However, animal materials, which are equally important in the TCM practice, are not well-known in other countries. On the other hand, the Chinese doctors had documented the toxic profiles of hundreds of animals and plants thousand years ago. Furthermore, they saw the potential benefits of these materials and used their toxic properties to treat a wide variety of diseases, such as heavy pain and cancer. Since the 50s of the last century, efforts of the Chinese government and societies to modernize TCM have achieved tremendous scientific results in both laboratory and clinic. A number of toxic proteins have been isolated and their functions identified. Although most of the literature was written in Chinese, this review provide a summary, in English, regarding our knowledge of the clinical use of the toxic proteins isolated from a plant, Tian Hua Fen, and an animal, scorpion, both of which are famous toxic prescriptions in TCM.

Download Full-text