scholarly journals Evaluation of five regions as DNA barcodes for identification of Lepista species (Tricholomataceae, Basidiomycota) from China

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e7307
Author(s):  
Siyu Wang ◽  
Hongbo Guo ◽  
JiaJia Li ◽  
Wei Li ◽  
Qin Wang ◽  
...  
Keyword(s):  
Ribosomal Rna ◽  
Internal Transcribed Spacer ◽  
Dna Barcode ◽  
Pcr Amplification ◽  
Intergenic Spacer ◽  
Its Region ◽  
Small Subunit ◽  
Dna Barcodes ◽  
Success Rates ◽  
Small Subunit Rdna

Background Distinguishing among species in the genus Lepista is difficult because of their similar morphologies. Methods To identify a suitable DNA barcode for identification of Lepista species, we assessed the following five regions: internal transcribed spacer (ITS), the intergenic spacer (IGS), nuclear ribosomal RNA subunit, mitochondrial small subunit rDNA, and tef1. A total of 134 sequences from 34 samples belong to eight Lepista species were analyzed. The utility of each region as a DNA barcode was assessed based on the success rates of its PCR amplification and sequencing, and on its intra- and inter-specific variations. Results The results indicated that the ITS region could distinguish all species tested. We therefore propose that the ITS region can be used as a DNA barcode for the genus Lepista. In addition, a phylogenetic tree based on the ITS region showed that the tested eight Lepista species, including two unrecognized species, formed eight separate and well-supported clades.

scholarly journals First Report of Phomopsis Stem Canker of Sunflower in Illinois Caused by Phomopsis helianthi

Plant Disease ◽  
2009 ◽  
Vol 93 (7) ◽  
pp. 760-760 ◽  
Author(s):  
M. D. Meyer ◽  
G. R. Zhang ◽  
D. K. Pedersen ◽  
C. A. Bradley
Keyword(s):  
Pcr Amplification ◽  
Its Region ◽  
Stem Canker ◽  
Small Subunit ◽  
The State ◽  
Lesion Length ◽  
Growth Stages ◽  
First Report ◽  
Small Subunit Rdna ◽  
Sunflower Production

Stem cankers were observed on confection sunflower (Helianthus annuus) plants growing in a field in Champaign County, Illinois in August 2008. Lesions were brown to reddish brown, elongated (approximately 10 to 15 cm long), and centered over the area where leaf petioles connected to the stems. Stem tissues underneath the lesions were degraded. Lesions from diseased stems were cut into 5- to 7-mm pieces and immersed in a 0.5% NaOCl solution for 1 min, rinsed with sterilized distilled water, and placed into petri dishes containing acidified potato dextrose agar (APDA; 4 ml of 25% lactic acid per liter). Fungal colonies that grew from the stem lesion pieces on APDA were white, floccose, and dense with dark colored substrate mycelia. On the basis of the symptoms on sunflower plants and the growth characteristics on APDA, the fungus was tentatively identified as Phomopsis helianthi (1). To confirm the identity of the fungus, PCR amplification of the small subunit rDNA and internal transcribed spacer (ITS) region with primers EF3RCNL and ITS4 was done (2). The PCR product was sequenced with these primers at the Keck Biotechnology Center at the University of Illinois, Urbana. The resulting nucleotide sequence was compared with small subunit rDNA and ITS sequences deposited in the nucleotide database ( http://www.ncbi.nlm.nih.gov ) and showed highest homology to sequences of Diaporthe helianthi, teleomorph of P. helianthi. To confirm pathogenicity of the fungus, sunflower plants (cv. Cargill 270) were grown in the greenhouse and inoculated with the isolated fungus. The stems of sunflower plants between the V2 and V4 growth stages (3) were excised just below the uppermost node. Mycelia plugs of the fungus were placed into the large end of disposable micropipette tips (200 μl). The micropipette tip containing the fungus was subsequently placed over a cut sunflower stem. The fungal isolate was used to inoculate five stems. To serve as controls, five cut sunflower stems were inoculated with micropipette tips containing plugs of noninfested PDA and five cut stems were not inoculated. Mean lesion length on the stem was measured from the inoculated tip toward the soil line 7 days after inoculation. The experiment was replicated over time. Mean lesion length over both replications averaged 24 mm on the fungus-inoculated plants, 2 mm on the noninfested PDA-inoculated control plants, and no lesions were present on the noninoculated control plants. The fungus was reisolated on PDA from the inoculated plants in the greenhouse. To our knowledge, this is the first report of P. helianthi causing a stem canker of sunflower in Illinois. Although commercial sunflower production in Illinois is currently limited, it is being evaluated as a potential crop to follow winter wheat in portions of the state. If sunflower production were to increase in the state, growers may have to monitor for and manage Phomopsis stem canker. References: (1) T. Gulya et al. Sunflower diseases. Page 263 in: Sunflower Technology and Production. American Society of Agronomy, Madison, WI, 1997. (2) N. S. Lord et al. FEMS Microbiol. Ecol. 42:327, 2002. (3) A. A. Schneiter and J. F. Miller, Crop Sci. 21:901, 1981.

Testing three proposed DNA barcodes for the wood identification of Dalbergia odorifera T. Chen and Dalbergia tonkinensis Prain

Holzforschung ◽  
2016 ◽  
Vol 70 (2) ◽  
pp. 127-136 ◽  
Author(s):  
Min Yu ◽  
Kai Liu ◽  
Liang Zhou ◽  
Lei Zhao ◽  
Shengquan Liu
Keyword(s):  
Economic Value ◽  
Dna Barcode ◽  
Pcr Amplification ◽  
First Grade ◽  
Wood Identification ◽  
Dna Barcodes ◽  
Success Rates ◽  
Interspecific Differences ◽  
Dalbergia Odorifera ◽  
Dalbergia Tonkinensis

Abstract Dalbergia odorifera T. Chen is a first-grade state protected plant in China. However, it is difficult to distinguish it from the closely related species Dalbergia tonkinensis Prain, which is less important in economic value, by wood anatomical features. In this study, three potential DNA barcode sequences, namely rpoC1, trnH-psbA and internal transcribed spacer (ITS), were used to differentiate wood of D. odorifera from D. tonkinensis. The average quantities of DNA extracts from twigs, sapwood and heartwood were 16.3, 11.5 and 6.0 ng mg-1, respectively. The success rates for polymerase chain reaction (PCR) amplification for three loci, namely ITS, trnH-psbA and rpoC1, were 62.5, 100 and 81.25%, respectively. The success rate for bidirectional sequencing of amplified products was 100% for all the three loci. The identification power of the three proposed DNA barcodes has been calculated by the BLAST, tree-based method and the TAXONDNA method. The interspecific differences of the trnH-psbA region were greater than intraspecific variations. Moreover, the identification power of trnH-psbA was higher than that of ITS and rpoC1 regions at the species level. Finally, the trnH-psbA region is proposed as a DNA barcode for wood identification between D. odorifera and D. tonkinensis.

scholarly journals First Report of Dollar Spot of Sandbur Caused by Sclerotinia homoeocarpa in Oklahoma

Plant Disease ◽  
2014 ◽  
Vol 98 (8) ◽  
pp. 1160-1160
Author(s):  
F. Flores ◽  
N. R. Walker
Keyword(s):  
Warm Season ◽  
Its Region ◽  
Small Subunit ◽  
Type I ◽  
Academic Press ◽  
Sclerotinia Homoeocarpa ◽  
Dollar Spot ◽  
First Report ◽  
Small Subunit Rdna ◽  
Disturbed Soils

Sandbur (Cenchrus incertus Curtis) is a warm-season, annual, noxious, grassy weed native to southern North America. It is common in sandy, disturbed soils and can also be found in home lawns and sport fields where low turf density facilitates its establishment. In July 2013, after a period of frequent rainfall and heavy dew, symptoms of dollar spot-like lesions (1) were observed on sandbur plants growing in a mixed stand of turf-type and native warm-season grasses in Logan County, Oklahoma. Lesions, frequently associated with leaf sheaths, were tan and surrounded by a dark margin. Symptomatic leaves were surface sterilized and plated on potato dextrose agar amended with 10 ppm rifampicin, 250 ppm ampicillin, and 5 ppm fenpropathrin. After incubation, a fungus morphologically identical to Sclerotinia homoeocarpa Bennett was consistently isolated. The nuclear ribosomal internal transcribed spacer (ITS) region of two different isolates, SCL2 and SCL3, were amplified using primers ITS4 and ITS5 (2). The DNA products were sequenced and BLAST analyses were used to compare sequences with those in GenBank. The sequence for isolate SLC2 was 869 bp, contained a type I intron in the 18S small subunit rDNA, and was identical to accession EU123803. The ITS sequence for isolate SLC3 was 535 bp and identical to accession EU123802. Twenty-five-day-old seedlings of C. incertus were inoculated by placing 5-mm-diameter agar plugs, colonized by mycelia of each S. homoeocarpa isolate, onto two of the plants' leaves. Plugs were held in place with Parafilm. Two plants were inoculated with each isolate and sterile agar plugs were placed on two leaves of another seedling as control. Plants were incubated in a dew chamber at 20°C and a 12-h photoperiod. After 3 days of incubation, water-soaked lesions surrounded by a dark margin appeared on inoculated plants only. Fungi that were later identified as S. homoeocarpa isolates SLC2 and SLC3 by sequencing of the ITS region were re-isolated from symptomatic leaves, fulfilling Koch's postulates. To our knowledge, this is the first report of dollar spot on sandbur. References: (1) R. W. Smiley et al. Page 22 in: Compendium of Turfgrass Diseases. 3rd ed. The American Phytopathological Society, St. Paul, MN, 2005. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

scholarly journals A Systematic Study on DNA Barcoding of Medicinally Important Genus Epimedium L. (Berberidaceae)

Genes ◽  
2018 ◽  
Vol 9 (12) ◽  
pp. 637 ◽  
Author(s):  
Mengyue Guo ◽  
Yanqin Xu ◽  
Li Ren ◽  
Shunzhi He ◽  
and Xiaohui Pang
Keyword(s):  
Dna Barcode ◽  
Morphological Characteristics ◽  
Identification Accuracy ◽  
Dna Barcodes ◽  
Success Rates ◽  
Data Set ◽  
Herba Epimedii ◽  
Potential Risks ◽  
Medicinal Properties ◽  
Species Being

Genus Epimedium consists of approximately 50 species in China, and more than half of them possess medicinal properties. The high similarity of species’ morphological characteristics complicates the identification accuracy, leading to potential risks in herbal efficacy and medical safety. In this study, we tested the applicability of four single loci, namely, rbcL, psbA-trnH, internal transcribed spacer (ITS), and ITS2, and their combinations as DNA barcodes to identify 37 Epimedium species on the basis of the analyses, including the success rates of PCR amplifications and sequencing, specific genetic divergence, distance-based method, and character-based method. Among them, character-based method showed the best applicability for identifying Epimedium species. As for the DNA barcodes, psbA-trnH showed the best performance among the four single loci with nine species being correctly differentiated. Moreover, psbA-trnH + ITS and psbA-trnH + ITS + rbcL exhibited the highest identification ability among all the multilocus combinations, and 17 species, of which 12 are medicinally used, could be efficiently discriminated. The DNA barcode data set developed in our study contributes valuable information to Chinese resources of Epimedium. It provides a new means for discrimination of the species within this medicinally important genus, thus guaranteeing correct and safe usage of Herba Epimedii.

The Low Copy Nuclear Gene Region, Granule Bound Starch Synthase (GBSS1), as a Novel Mini-DNA Barcode for the Identification of Different Sage (Salvia) Species

Planta Medica ◽  
2021 ◽  
Author(s):  
Iffat Parveen ◽  
Natascha Techen ◽  
Sara M. Handy ◽  
Jing Li ◽  
Charles Wu ◽  
...  
Keyword(s):  
Internal Transcribed Spacer ◽  
Dna Barcode ◽  
Nuclear Genome ◽  
Nuclear Gene ◽  
Starch Synthase ◽  
Species Discrimination ◽  
Success Rates ◽  
Discrimination Ability ◽  
Internal Transcribed Spacer 2 ◽  
Granule Bound Starch Synthase

AbstractMorphological similarity within species makes the identification and authentication of Salvia species challenging, especially in dietary supplements that contain processed root or leaf powder of different sage species. In the present study, the species discriminatory power of 2 potential DNA barcode regions from the nuclear genome was evaluated in 7 medicinally important Salvia species from the family Lamiaceae. The nuclear internal transcribed spacer 2 and the exon 9 – 14 region of low copy nuclear gene WAXY coding for granule-bound starch synthase 1 were tested for their species discrimination ability using distance, phylogenetic, and BLAST-based methods. A novel 2-step PCR method with 2 different annealing temperatures was developed to achieve maximum amplification from genomic DNA. The granule-bound starch synthase 1 region showed higher amplification and sequencing success rates, higher interspecific distances, and a perfect barcode gap for the tested species compared to the nuclear internal transcribed spacer 2. Hence, these novel mini-barcodes generated from low copy nuclear gene regions (granule-bound starch synthase) that were proven to be effective barcodes for identifying 7 Salvia species have potential for identification and authentication of other Salvia species.

scholarly journals Five Fungal Pathogens Are Responsible for Bayberry Twig Blight and Fungicides Were Screened for Disease Control

2020 ◽  
Vol 8 (5) ◽  
pp. 689 ◽  
Author(s):  
Wenjun Li ◽  
Ming Hu ◽  
Yang Xue ◽  
Zhijun Li ◽  
Yanfei Zhang ◽  
...  
Keyword(s):  
Ribosomal Rna ◽  
Fungal Pathogens ◽  
Large Subunit ◽  
Elongation Factor ◽  
Morphological Characteristics ◽  
Its Region ◽  
Small Subunit ◽  
Field Application ◽  
Financial Loss ◽  
Twig Blight

Bayberry (Myrica rubra) is a commercial fruit in China. For the past seven years, twig blight disease has been attacking bayberry plantations in Shantou City, Guangdong Province, China, leading to destructive damage and financial loss. In this study, five fungal species associated with twig dieback and stem blight were identified based on morphological characteristics combined with multilocus sequence analysis (MLSA) on the internal transcribed spacer (ITS) region, partial sequences of β-tubulin (tub2), translation elongation factor 1-α (tef1-α), large subunit ribosomal RNA (LSU) and small subunit ribosomal RNA (SSU) genes, which are Epicoccum sorghinum, Neofusicoccum parvum, Lasiodiplodia theobromae, Nigrospora oryzae and a Pestalotiopsis new species P. myricae. P. myricae is the chief pathogen in fields, based on its high isolation rate and fast disease progression after inoculation. To our knowledge, this is the first study reporting the above five fungi as the pathogens responsible for bayberry twig blight. Indoor screening of fungicides indicates that Prochloraz (copper salt) is the most promising fungicide for field application, followed by Pyraclostrobin, 15% Difenoconazole + 15% Propiconazole, Difenoconazole and Myclobutanil. Additionally, Bacillus velezensis strain 3–10 and zeamines from Dickeya zeae strain EC1 could be used as potential ecofriendly alternatives to control the disease.

scholarly journals An ITS-based phylogenetic framework for the genus Vorticella : finding the molecular and morphological gaps in a taxonomically difficult group

2013 ◽  
Vol 280 (1771) ◽  
pp. 20131177 ◽  
Author(s):  
Ping Sun ◽  
John C. Clamp ◽  
Dapeng Xu ◽  
Bangqin Huang ◽  
Mann Kyoon Shin ◽  
...  
Keyword(s):  
Ribosomal Rna ◽  
Phylogenetic Relationships ◽  
Internal Transcribed Spacer ◽  
Small Subunit ◽  
Rrna Gene ◽  
Ssu Rrna ◽  
Branching Order ◽  
Close Relationship ◽  
Molecular Phylogenetic ◽  
Phylogenetic Framework

Vorticella includes more than 100 currently recognized species and represents one of the most taxonomically challenging genera of ciliates. Molecular phylogenetic analysis of Vorticella has been performed so far with only sequences coding for small subunit ribosomal RNA (SSU rRNA); only a few of its species have been investigated using other genetic markers owing to a lack of similar sequences for comparison. Consequently, phylogenetic relationships within the genus remain unclear, and molecular discrimination between morphospecies is often difficult because most regions of the SSU rRNA gene are too highly conserved to be helpful. In this paper, we move molecular systematics for this group of ciliates to the infrageneric level by sequencing additional molecular markers—fast-evolving internal transcribed spacer (ITS) regions—in a broad sample of 66 individual samples of 28 morphospecies of Vorticella collected from Asia, North America and Europe. Our phylogenies all featured two strongly supported, highly divergent, paraphyletic clades (I, II) comprising the morphologically defined genus Vorticella . Three major lineages made up clade I, with a relatively well-resolved branching order in each one. The marked divergence of clade II from clade I confirms that the former should be recognized as a separate taxonomic unit as indicated by SSU rRNA phylogenies. We made the first attempt to elucidate relationships between species in clade II using both morphological and multi-gene approaches, and our data supported a close relationship between some morphospecies of Vorticella and Opisthonecta , indicating that relationships between species in the clade are far more complex than would be expected from their morphology. Different patterns of helix III of ITS2 secondary structure were clearly specific to clades and subclades of Vorticella and, therefore, may prove useful for resolving phylogenetic relationships in other groups of ciliates.

scholarly journals Nuclear ribosomal internal transcribed spacer (ITS) region as a universal DNA barcode marker for Fungi

2012 ◽  
Vol 109 (16) ◽  
pp. 6241-6246 ◽  
Author(s):  
C. L. Schoch ◽  
K. A. Seifert ◽  
S. Huhndorf ◽  
V. Robert ◽  
J. L. Spouge ◽  
...  
Keyword(s):  
Internal Transcribed Spacer ◽  
Dna Barcode ◽  
Its Region

scholarly journals Detection of Pythium ultimum Using Polymerase Chain Reaction with Species-Specific Primers

Plant Disease ◽  
1997 ◽  
Vol 81 (10) ◽  
pp. 1155-1160 ◽  
Author(s):  
K. Kageyama ◽  
A. Ohyama ◽  
M. Hyakumachi
Keyword(s):  
Polymerase Chain Reaction ◽  
Internal Transcribed Spacer ◽  
Pcr Amplification ◽  
Its Region ◽  
Pythium Ultimum ◽  
Pythium Species ◽  
Chain Reaction ◽  
Specific Primers ◽  
Polymerase Chain ◽  
Species Specific

This study was conducted to sequence the rDNA internal transcribed spacer (ITS) region of Pythium ultimum and Pythium group HS, design species-specific primers for polymerase chain reaction (PCR), and detect P. ultimum from diseased seedlings using PCR. The sequence of the ITS region of P. ultimum was identical with that of Pythium group HS. The results support the reports that the HS group is an asexual strain of P. ultimum. Using PCR, the primer pair K1+K3, designed on portions of the sequence of the ITS region, amplified isolates of P. ultimum and the HS group but not isolates of 20 other Pythium species. DNA extracts from damped-off seedlings were not amplified, but a 10-fold dilution of the extracts with Tris-EDTA (TE) buffer diluted the inhibitors and allowed PCR amplification. The primer pair used detected P. ultimum from a single diseased seedling.

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