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

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.

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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.

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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.

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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.

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Development of Race-Specific SCAR Markers for Detection of Chinese Races CYR32 and CYR33 of Puccinia striiformis f. sp. tritici

Plant Disease ◽

10.1094/pdis-94-2-0221 ◽

2010 ◽

Vol 94 (2) ◽

pp. 221-228 ◽

Cited By ~ 7

Author(s):

Baotong Wang ◽

Xiaoping Hu ◽

Qiang Li ◽

Baojun Hao ◽

Bo Zhang ◽

...

Keyword(s):

Molecular Markers ◽

Stripe Rust ◽

Genomic Dna ◽

Puccinia Striiformis ◽

Random Amplified Polymorphic Dna ◽

Scar Markers ◽

Wheat Stripe Rust ◽

Wheat Genotypes ◽

Sequence Characterized Amplified Region ◽

Wheat Leaves

Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici, is a devastating disease in China. Races CYR32 and CYR33 have been predominant in the recent P. striiformis f. sp. tritici population. To develop molecular markers for these races, initially 86 isolates, most of which were collected in 2007 throughout China, were tested on the set of wheat genotypes for differentiating Chinese P. striiformis f. sp. tritici races, and their genomic DNA were amplified with 94 random amplified polymorphic DNA (RAPD) primers. Twelve isolates were identified as CYR33, 14 as CYR32, and 60 as 13 other races. A 320-bp band was identified to be associated with CYR32 with primer S1271 (5′-CTTCTCGGTC-3′), and a 550-bp band was identified to be specific to CYR33 with primer S1304 (5′-AGGAGCGACA-3′). The two bands were cloned and sequenced. Based on the sequences, sequence characterized amplified region (SCAR) markers CYR32sp1/sp2 and CYR33sp1/sp2 were developed to differentiate CYR32 and CYR33, respectively, from other races. The SCAR markers were validated with DNA samples from wheat leaves inoculated with selected isolates from the 86 isolates and urediniospore DNA samples from an additional 63 isolates collected from 2006 to 2009. The detection of CYR32 and CYR33 with the SCAR markers was completely consistent with the results of the race identification with the set of differential wheat genotypes. Thus, the markers are highly reliable for identification of the two races.

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

Molecules ◽

10.3390/molecules23071748 ◽

2018 ◽

Vol 23 (7) ◽

pp. 1748 ◽

Cited By ~ 8

Author(s):

Inkyu Park ◽

Sungyu Yang ◽

Wook Kim ◽

Pureum Noh ◽

Hyun Lee ◽

...

Keyword(s):

Phylogenetic Analysis ◽

Scar Marker ◽

Herbal Medicines ◽

The Other ◽

Evolutionary Information ◽

Scar Markers ◽

Dna Barcodes ◽

Genomic Information ◽

Sequence Characterized Amplified Region ◽

Dipsacus Asper

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.

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SCAR Marker for Gender Identification in Date Palm (Phoenix dactylifera L.) at the Seedling Stage

International Journal of Genomics ◽

10.1155/2018/3035406 ◽

2018 ◽

Vol 2018 ◽

pp. 1-6 ◽

Cited By ~ 1

Author(s):

Fahad Al-Qurainy ◽

Abdulhafed A. Al-Ameri ◽

Salim Khan ◽

Mohammad Nadeem ◽

Abdel-Rhman Z. Gaafar ◽

...

Keyword(s):

Date Palm ◽

Scar Marker ◽

Phoenix Dactylifera ◽

Seedling Stage ◽

Gender Identification ◽

Specific Primers ◽

Sequence Characterized Amplified Region ◽

Pcr Product ◽

Phoenix Dactylifera L ◽

Reproducible Band

Date palm (Phoenix dactylifera L.) is cultivated in arid and semiarid regions worldwide. Given the dioecious nature of this plant, gender identification is very important at the seedling stage. Molecular markers are very effective tools that help in gender identification at this stage. A sequence characterized amplified region (SCAR) marker linked to sex-specific regions in the genome of date palm was developed. Of the 300 tested randomly amplified polymorphic DNA (RAPD) primers, only one primer (OPC-06) produced reproducible band (294 bp) in male plants. The PCR product of this primer was cloned and sequenced. The specific primers were synthesized for amplification of a 186 bp fragment in male date palm plants. These primers were validated in male and female date palm plants, wherein the designed SCAR marker was reported only in male plants and no amplification was observed in female plants. The developed SCAR marker was used with seedlings of date palm and proved very effective in identification of gender.

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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.

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Comparison of Anthracnose Resistance with the Presence of Two SCAR Markers Associated with the Rca2 Gene in Strawberry

HortScience ◽

10.21273/hortsci13805-18 ◽

2019 ◽

Vol 54 (5) ◽

pp. 793-798 ◽

Cited By ~ 1

Author(s):

Melinda A. Miller-Butler ◽

Barbara J. Smith ◽

Brian R. Kreiser ◽

Eugene K. Blythe

Keyword(s):

Molecular Markers ◽

Agronomic Traits ◽

Scar Markers ◽

Breeding Programs ◽

Anthracnose Resistance ◽

Sequence Characterized Amplified Region ◽

Starting Point ◽

Strawberry Anthracnose ◽

Strawberry Cultivars ◽

Breeding Germplasm

Strawberry anthracnose diseases are caused primarily by three Colletotrichum species: C. acutatum J.H. Simmonds, C. fragariae A.N. Brooks, and C. gloeosporioides (Penz.) Penz. & Sacc. Molecular markers are being used in breeding programs to identify alleles linked to disease resistance and other positive agronomic traits. In our study, strawberry cultivars and breeding germplasm with known anthracnose susceptibility or resistance to the three anthracnose-causing Colletotrichum species were screened for two sequence characterized amplified region (SCAR) markers linked to the Rca2 gene. The Rca2 resistant allele SCAR markers were associated with varying degrees of significance for a strawberry plant’s anthracnose resistance to C. fragariae but not to C. acutatum or C. gloeosporioides. Although the presence or absence of the markers associated with the Rca2 resistance gene is an imperfect indicator of anthracnose resistance, it may serve as a useful starting point in selecting germplasm for breeding programs.

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Species-diagnostic and species-specific DNA sequences evenly distributed throughout pine and spruce chromosomes

Genome ◽

10.1139/g10-065 ◽

2010 ◽

Vol 53 (10) ◽

pp. 769-777 ◽

Cited By ~ 1

Author(s):

Melanie Mehes-Smith ◽

Paul Michael ◽

Kabwe Nkongolo

Keyword(s):

Dna Sequences ◽

Random Amplified Polymorphic Dna ◽

Inter Simple Sequence Repeat ◽

Issr Markers ◽

Pinus Monticola ◽

The Family ◽

Species Specific ◽

Rapd And Issr ◽

Specific Sequences

Genome organization in the family Pinaceae is complex and largely unknown. The main purpose of the present study was to develop and physically map species-diagnostic and species-specific molecular markers in pine and spruce. Five RAPD (random amplified polymorphic DNA) and one ISSR (inter-simple sequence repeat) species-diagnostic or species-specific markers for Picea mariana , Picea rubens , Pinus strobus , or Pinus monticola were identified, cloned, and sequenced. In situ hybridization of these sequences to spruce and pine chromosomes showed the sequences to be present in high copy number and evenly distributed throughout the genome. The analysis of centromeric and telomeric regions revealed the absence of significant clustering of species-diagnostic and species-specific sequences in all the chromosomes of the four species studied. Both RAPD and ISSR markers showed similar patterns.

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