• Proceedings of the Korean Society of Plant Pathology Conference
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• 2003.10a
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• pp.131.1-131
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• 2003

This study reports the identification of species of Colletotrichum strains originating from red-pepper and Chinese matrimony vine in Cheongyang. Ninteen isolates of red-pepper and 26 Coiletotrichum isolates of Chinese matrimony vine were compared with 5 isolates of strawberry representing C. gloeosporioides, by use of morphological and cultural criteria. Twenty three isolates among 26 isolates from Chinese matrimony vine were identified as C. acutatum, characterized by the low growth rates and the low sensitivity to carbendazim and diethofencarb. Also, all the isolates of red-pepper were identified as C. acutatum, showing the same characteristics as those of Chinese matrimony vine. Three and five isolates from Chinese matrimony vine and strawberry, respectively, were identified as C. gloeosporioides, characterized by the high growth rates and the high seneitivity to carbendazim and diethofencarb. There were differences in colony color and pathogenicity between Chinese matrimony vine isolates and red-pepper isolates of C. autatum. The isolates of C. acutatum from Chinese matrimony vine producing orange colored colonies with abundant spores showed the strong pathogenicity to Chinese matrimony vine, although they could not infect fruits of red-pepper by the wound inoculation. However, red-pepper isolates of C. acutatum producing gray colonies showed the strong pathogenicity to Chinese matrimony vine as well as red-pepper. Furthermore, comparative study on PCR amplification of ITS regions of rDNA was carried out using a number of Colletotrichum isolates. A species-specific primer could be used for the identification of C. acutatum from red-pepper and Chinese matrimony vine.

• Korean Journal of Medicinal Crop Science
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• v.26 no.4
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• pp.302-307
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• 2018

Background: A soil-borne pathogenic fungus, Ilyonectria radicicola (Cylindrocarpon destructans) causes root rot on ginseng (Panax ginseng C. A. Meyer) and is known to attack many other plants. The Nectria/Neonectria radicicola complex has been renamed as the I. radicicola complex after analysis of its multi-gene relatedness and morphological characteristics. The fungi in this complex have been reclassified into 16 species under the genus Ilyonectria based on characteristics analysis Methods and Results: To obtain useful data from the Korean ginseng root rot, I. radicicola was isolated from the rhizosphere soils of the chestnut tree. They were identified through a pathogenicity test and a survey of the morphological features. The existence of I. radicicola in soil samples was confirmed by PCR detections using nested PCR with species-specific primer sets. These were subsequenctly isolated on semi-selective media from PCR-positive soils. Genetic analysis of the I. radicicola complex containing these pathogens was done by comparing the DNA sequences of the histone h3 region. These isolates originating from the rhizosphere soils of chestnut constituted a clade with other closely related species or I. radicicola isolates originating from ginseng or other host plants, respectively. Additionally, the pathogenicity tests to analyze the characteristics of these I. radicicola isolates revealed that they caused weakly virulent root rot on ginseng. Conclusions: This is the first study reporting that I. radicicola isolates from chestnut rhizosphere soils can attack ginseng plant in Korea. Thus, these results are expected to provide informations in the selection of suitable fields for ginseng cultivation.

• Korean Journal of Plant Resources
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• v.14 no.3
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• pp.235-240
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• 2001

Polymerase chain reaction (PCR) is used in a wide array of researches in plant molecular genetics and breeding. However, considerable time and cost are still required for the preparation of DNA suitable for reliable PCR results, especially in plant species containing high amounts of polyphenols. To reduce time and effort for PCR-based analysis, a simplified but reliable method was developed by a combinational employment of a simple and fast DNA extraction procedure and BLOTTO (Bovine Lacto Transfer Technique Optimizer) in reaction mixture. Genomic DNAs prepared by one-step extraction method from recalcitrant plant species such as Rubus coreanus, apple, grape and lettuce were successfully amplified by random primers in the reaction mixture containing 2 to 4% BLOTTO. Successful amplification of ${\gamma}$-TMT transgene in lettuce transformants by the specific primers was also achieved in the same condition, making rapid screening of positive transformants possible. Our results suggest that use of a simple DNA extraction procedure and incorporation of BLOTTO in reaction mixture in combination can reduce time and effort required for the analyses of a large number of germplasms and transformants by PCR-based techniques.

• Horticultural Science & Technology
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• v.33 no.2
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• pp.242-250
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• 2015

Insertional mutagenesis induced by T-DNA or transposon tagging offers possibilities for analysis of gene function. However, its potential remains limited unless good methods for detecting the target locus are developed. We describe a PCR technique for efficient identification of DNA sequences adjacent to the inserted T-DNA in a higher plant, Chinese cabbage (Brassica rapa ssp. pekinensis). This strategy, which we named variable argument thermal asymmetric interlaced PCR (VA-TAIL PCR), was designed by modifying a single-step annealing-extension PCR by including a touch-up PCR protocol and using long gene-specific primers. Amplification efficiency of this PCR program was significantly increased by employing an autosegment extension method and linked sequence strategy in nested long gene-specific primers. For this technique, arbitrary degenerate (AD) primers specific to B. rapa were designed by analyzing the Integr8 proteome database. These primers showed higher accuracy and utility in the identification of flanking DNA sequences from individual transgenic Chinese cabbages in a large T-DNA inserted population. The VA-TAIL PCR method described in this study allows the identification of DNA regions flanking known DNA fragments. This method has potential biotechnological applications, being highly suitable for identification of target genomic loci in insertional mutagenesis screens.

• Journal of Food Hygiene and Safety
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• v.33 no.4
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• pp.306-309
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• 2018

The present study was conducted to investigate the prevalence of Mycoplasma spp. in cows and pigs with pneumonic lung lesions in Gyeonggi province in 2017. One hundred ninety two and 257 lung tissues were collected from slaughtered cows and pigs with pneumonic lesions, respectively, and examined for the presence of Mycoplasma spp. by a genus specific PCR. Among the examined animals, 147 cows (76.5%) and 203 pigs (80.9%) were found to be infected with Mycoplasma spp.. The infected tissues were further examined to identify the specific species of Mycoplasma using species specific PCRs. The only identified species in cows was M. agalactiae which was detected from 16 cows (8.3%), whereas M. dispar, M. bovis, and M. bovirhinis were not detected. In pigs, M. hyopneumoniae was detected from 74 pigs (28.8%) and M. hyorhinis from 13 pigs (5.1%). M. hyosynoviae was not detected. Taken together, the current study indicates Mycoplasma spp. are commonly associated with lung infection in cows and pigs in Korea. Further studies are needed to evaluate the impact of mycoplasma infection on the development of lung diseases in farm animals.

• Journal of Food Hygiene and Safety
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• v.35 no.3
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• pp.234-242
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• 2020

In this study, we monitored the raw materials in home-meal replacement (HMR) products, which have shown more than 63% growth in market size for two years. A total of 89 HMR products were purchased and the DNA barcodes of 112 raw materials in the product samples were analyzed. In order to identify the raw material species, a primer set specific for the 16S ribosomal RNA region of each raw material species was amplified. The amplicon was purified and sequenced, and then used to perform a BLAST search provided by the National Institutes of Health (NIH). The species of the raw material was determined by comparing the nucleotide sequences of the species registered in GenBank with identity and match score. Twenty-four species and three genera were identified from 112 raw materials. Three genera were identified at the genus level because a large number of species belonging to the same genus exist within 98% of the identity criteria. The results of the determination were compared with the available raw materials suggested in the Korea Food Code to determine the Korean name and availability of the foods. Six non-listed species were determined to be edible according to information provided by influential domestic and foreign organizations.

• The Korean Journal of Mycology
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• v.39 no.1
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• pp.31-38
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• 2011

In this study, 81 commercial strains of Pleurotus species cultivated in South Korea were analyzed with randomly amplified polymorphic DNA (RAPD) technique. Sequence characterized amplified region (SCAR) markers were developed by designing from one RAPD polymorhic band specific to Suhan strain. The SCAR primer pair 'S-OPA13-1' amplified a 590-bp fragment in the varieties originated from Suhan strain. The Blast search of S-OPA13-1 showed high homology to the POMFBO1 P. ostreatus cDNA clone MFB02-A05 and Laccaria bicolor S238N-H82. The results showed that this SCAR marker can clearly distinguish Suhan strains from Pleurotus spp.

• Journal of Microbiology and Biotechnology
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• v.15 no.3
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• pp.502-509
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• 2005

Sexual reproduction plays an important role in the biology and epidemiology of oomycete plant pathogens such as the heterothallic species Phytophthora infestans. Recent worldwide dispersal of A2 mating type strains of P. infestans resulted in increased virulence, gene transfer, and genetic variation, creating new challenges for disease management. To develop a genetic assay for mating type identification in P. infestans, we used the Amplified Fragment Length Polymorphism (AFLP) technique. The primer combination E+AT/M+CTA detected a fragment specific to A1 mating type (Mat-A1) of P. infestans. This fragment was cloned and sequenced, and a pair of primers (INF-1, INF-2) were designed and used to differentiate P. infestans Mat-A1 from Mat-A2 strains. The Mat A1-specific fragment was detected using Southern blot analysis of PCR products amplified with primers INF-1 and INF-2 from genomic DNA of 14 P. infestans Mat-A1 strains, but not 13 P. infestans Mat-A2 strains or 8 other isolates representing several Phytophthora spp. Southern blot analysis of genomic DNAs of P. infestans isolates revealed a 1.6 kb restriction enzyme (EcoRI, BamHI, AvaI)-fragment only in Mat-A1 strains. The A1 mating type-specific primers amplified a unique band under stringent annealing temperatures of $63^{\circ}C-64^{\circ}C$, suggesting that this PCR assay could be developed into a useful method for mating type determination of P. infestans in field material.

• Mycobiology
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• v.41 no.2
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• pp.86-93
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• 2013

Sequence characterized amplified region (SCAR) markers are one of the most effective and accurate tools for microbial identification. In this study, we applied SCAR markers for the rapid and accurate detection of Phytophthora katsurae, the casual agent of chestnut ink disease in Korea. In this study, we developed seven SCAR markers specific to P. katsurae using random amplified polymorphic DNA (RAPD), and assessed the potential of the SCAR markers to serve as tools for identifying P. katsurae. Seven primer pairs (SOPC 1F/SOPC 1R, SOPC 1-1F/SOPC 1-1R, SOPC 3F/SOPC 3R, SOPC 4F/SOPC 4R, SOPC 4F/SOPC 4-1R, SOPD 9F/SOPD 9R, and SOPD 10F/SOPD 10R) from a sequence derived from RAPD fragments were designed for the analysis of the SCAR markers. To evaluate the specificity and sensitivity of the SCAR markers, the genomic DNA of P. katsurae was serially diluted 10-fold to final concentrations from 1 mg/mL to 1 pg/mL. The limit of detection using the SCAR markers ranged from $100{\mu}g/mL$ to 100 ng/mL. To identify the limit for detecting P. katsurae zoospores, each suspension of zoospores was serially diluted 10-fold to final concentrations from $10{\times}10^5$ to $10{\times}10^1$ zoospores/mL, and then extracted. The limit of detection by SCAR markers was approximately $10{\times}10^1$ zoospores/mL. PCR detection with SCAR markers was specific for P. katsurae, and did not produce any P. katsurae-specific PCR amplicons from 16 other Phytophthora species used as controls. This study shows that SCAR markers are a useful tool for the rapid and effective detection of P. katsurae.

• Food Science and Biotechnology
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• v.17 no.4
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• pp.761-765
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• 2008

Bacillus cereus causes two different types of food poisoning syndromes: diarrhea and emesis. The diarrheal syndrome is attributed to various enterotoxins, including nonhemolytic enterotoxin, hemolytic enterotoxin, and enterotoxin-T, whereas the emetic syndrome is caused by the dodecadepsipeptide toxin cereulide. A multiplex polymerase chain reaction (PCR) assay was developed to rapidly detect and identify B. cereus strains. Three primer pairs specific to regions within genes encoding nonhemolytic enterotoxin (nheA), molecular chaperonin (groEL), and cereulide synthetase (ces) were used to identify and differentiate between the enterotoxin-producing and emetic toxin-producing B. cereus strains. The cereulide-producing emetic B. cereus showed 3 PCR products of 325, 405, and 685 bp for the groEL, ces, and nheA genes, respectively, whereas the enterotoxin-producing B. cereus showed 2 PCR products without a ces gene specific DNA fragment. Specific amplifications and differentiations by multiplex PCR assay were obtained using 62 B. cereus strains and 13 strains' of other bacterial species. The detection limit of this assay for enterotoxin-producing strain and emetic toxin-producing strain from pure cultures were $2.4{\times}10^1$ and $6.0{\times}10^2\;CFU/tube$, respectively. These results suggest that our multiplex PCR method may be useful for the rapid detection and differentiation of B. cereus strains in foods.