• Journal of Korean Society of Forest Science
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• v.104 no.2
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• pp.332-335
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• 2015

Taphrina wiesneri, a pathogen of cherry witches' broom, is highly pathogenic to Prunus yedoensis Matsumura which are widely planted in parks and streets in South Korea. In order to control the disease, it is crucial to know the life cycle of the fungus. We attempted to detect the fungus tentatively overwintering in shoots and branches of cherry trees both having witches' broom and healthy before flowering and leafing in spring using PCR with species-specific primer set (TwITSF and TwITSR). Genomic DNAs were extracted from the symptomatic and the asymptomatic shoots or branches. Results indicated that T. wiesneri is present in leaf buds and inner bark not only in symptomatic branches but also in the asymptomatic branches in diseased trees. However, the fungus was not detected in flower buds of the symptomatic trees and any samples of healthy trees.

• The Plant Pathology Journal
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• v.24 no.1
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• pp.17-23
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• 2008

A total of 33 isolates of Colletotrichum species obtained from pepper, apple, and strawberry in 2001 and 2002 were identified based on mycological characteristics, responses to fungicides carbendazim and the mixture of carbendazim and diethofencarb, and nucleotide sequence analysis of internal transcribed spacer (ITS) regionMost of the Colletotrichum isolates from pepper could be identified as C. acutatum. The pepper isolates produced grey white mycelia that gradually changed to dark gray. The conidia were variable in size, and almost cylindrical in shape with at least one rounded end. They could grow on PDA amended with carbendazim or with the mixture of carbendazim and diethofencarb at $10{\mu}g/ml$, to which the isolates from apple and strawberry were very sensitive. A part of the ITS regions from the Colletotrichum isolates was amplified with the specific primers designed for C. acutatum (Ca1-1) or C. gloeosporioides (Cg1-3). A primer pair of Ca1-1 and a universal primer (ITS4) amplified a 496-bp DNA fragment from all of the pepper isolates examined and one apple isolate. Taken together, it is conclusive that the Colletotrichum isolates causing the typical lesion of anthracnose on pepper fruits are C. acutatum.

• Journal of Ginseng Research
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• v.43 no.1
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• pp.1-9
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• 2019

Background: Korean ginseng is an important cash crop in Asian countries. However, plant yield is reduced by pathogens. Among the Ilyonectria radicicola-species complex, I. mors-panacis is responsible for root-rot and replant failure of ginseng in Asia. The development of new methods to reveal the existence of the pathogen before cultivation is started is essential. Therefore, a quantitative real-time polymerase chain reaction method was developed to detect and quantify the pathogen in ginseng soils. Methods: In this study, a species-specific histone H3 primer set was developed for the quantification of I. mors-panacis. The primer set was used on DNA from other microbes to evaluate its sensitivity and selectivity for I. mors-panacis DNA. Sterilized soil samples artificially infected with the pathogen at different concentrations were used to evaluate the ability of the primer set to detect the pathogen population in the soil DNA. Finally, the pathogen was quantified in many natural soil samples. Results: The designed primer set was found to be sensitive and selective for I. mors-panacis DNA. In artificially infected sterilized soil samples, using quantitative real-time polymerase chain reaction the estimated amount of template was positively correlated with the pathogen concentration in soil samples ($R^2=0.95$), disease severity index ($R^2=0.99$), and colony-forming units ($R^2=0.87$). In natural soils, the pathogen was recorded in most fields producing bad yields at a range of $5.82{\pm}2.35pg/g$ to $892.34{\pm}103.70pg/g$ of soil. Conclusion: According to these results, the proposed primer set is applicable for estimating soil quality before ginseng cultivation. This will contribute to disease management and crop protection in the future.

• Food Engineering Progress
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• v.21 no.2
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• pp.174-179
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• 2017

For preliminary molecular typing, PCR-based fingerprinting using random amplified polymorphic DNA (RAPD) is the method of choice. In this study, 14 bacterial strains were isolated from different Korean food sources, identified using 16S rRNA gene sequencing, and characterized through RAPD-PCR. Two PCR primers (239 and KAY3) generated a total of 130 RAPD bands, 14 distinct PCR profiles, 10 polymorphic bands, one monomorphic band, and four unique bands. Dendrogram-based analysis with primer 239 showed that all 14 strains could be divided into seven clades out of which clade VII had the maximum of seven. In contrast, dendrogram analysis with the primer KAY3 divided the 14 L. citreum strains into four clades out of which clade IV consisted of a maximum of 10 strains out of 14. This research identified and characterized bacterial populations associated with different Korean foods. The proposed RAPD-PCR method, based on sequence amplification, could easily identify and discriminate the lactic acid bacteria species at the strain-specific level and could be used as a highly reliable genomic fingerprinting tool.

• The Plant Pathology Journal
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• v.15 no.4
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• pp.228-235
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• 1999

Genetic diversity of ninety-five Korean isolates of Phytophthora was investigated on the basis of PCR-RFLP of ribosomal DNA. The isolates were previously identified as following fifteen species by mycological and cultural characteristics; P. boehmeriae, P. cactorum, P. cambivora, P. capsici, P. cinnamoni, P. citricola, P. citrophthora, P. cryptogea, P. drechsleri, P. erythroseptica, P. infestans, P. megasperma, P. nicotianae, P. palmivora and P. sojae. The regions of small subunit (SSU) and internal transcribed spacer (ITS) of rDNA were amplified with primer pair, NS1 and ITS4, by polymerase chain reaction (PCR) and digested with nine restriction enzymes. P. boehmeriae, P. cactorum, P. cambivora, P. capsici, P. cinnamomi, P. citricola, P. citrphthora, P. infestans, P. nicotianae and P. palmivora showed specific band patterns for each species. However, P. sojae and P. erythroseptica presented identical band patterns and P. cryptogea, P. drechsleri and P. megasperma were divided into six groups, which were not compatible with delineation of the species. A group originated from cucurbits showed distinct band patterns from other groups, but the other five groups were closely related within 96.0% similarity, forming one complex group. Consequently, Korean isolates of Phytophthora were divided into thirteen genetic groups and each group was readily differentiated by comparing digestion patterns of AvaII, HaeIII, MboI, HhaI and MspI. Therefore, PCR-RFLP of rDNA using the five enzymes can be used to differentiate or identify the Phytophthora species reported in Korea so far.

• Microbiology and Biotechnology Letters
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• v.36 no.2
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• pp.96-100
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• 2008

Diversity of lactic acid bacteria involved in 5 Korean fermented vegetables (Cot kimchi, Dongchimi, Baechu kimchi, Oisobagi, and Chonggak kimchi) was investigated using PCR-based method. PCR primer pairs targeted the recA gene were used for the detection of 7 species of lactic acid bacteria mainly found in kimchi and Lactobacillus acidophilus involved in dairy fermentation. Lactobacillus plantarum and Lactobacillus sakei were detected in all samples tested but Lactobacillus paraplantarum, Lactobacillus pentosus, and Lb. acidophilus were not detected. Lactobacillus brevis and Leuconostoc citreum were detected only from Baechu kimchi and Leuconostoc mesenteroides was detected from Got kimchi, Dongchimi, Baechu kimchi, and Oisobagi. The difference of detected species from fermented vegetables may be originated from the difference of main materials. Lb. plantarum and Lb. sakei are supposed to be broadly involved in Korean fermented vegetables.

• BMB Reports
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• v.37 no.2
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• pp.213-222
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• 2004

A randomly amplified polymorphic DNA (RAPD) analysis was used to identify the species- and population-specific markers of abalone; Haliotis asinina, H. ovina, and H. varia in Thai waters. Fifteen species-specific and six population-specific RAPD markers were identified. In addition, an 1650 bp band (UBC195) that was restricted to H. ovina from the Gulf of Thailand (east) was also found. All of the specific RAPD markers were cloned and sequenced. Twenty pairs of primers were designed and specificity-tested (N = 12 and 4 for target and non-target species, respectively). Seven primer pairs (CUHA1, 2, 4, 11, 12, 13, and 14) were specifically amplified by H. asinina DNA, whereas a single pair of primers showed specificity with H. ovina (CUHO3) and H. varia (CUHV1), respectively. Four primer pairs, including CUHA2, CUHA12, CUHO3, and CUHV1, were further examined against 216 individuals of abalone (N = 111, 73, and 32, respectively). Results indicated the species-specific nature of all of them, except CUHO3, with the sensitivity of detection of 100 pg and 20 pg of the target DNA template for CUHA2 and CUHA12 and CUHV1, respectively. The species-origin of the frozen, ethanol-preserved, dried, and boiled H. asinina specimens could also be successfully identified by CUHA2.

• Korean Journal of Plant Resources
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• v.17 no.2
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• pp.107-115
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• 2004

To develop universal primers for phylogenetic analysis and species-specific marker for breeding program of kiwifruit, eighteens primers were designed from kiwifruit genome-specific repeat sequences. Seven species including twenty two varieties collected from native eastern Asia were examined using 18 to 22 mer kiwifruit target(KT) primers. among eighteen primers, we selected seven primers for phylogenetic relationship. The genus Actinidia was divided into two large groups; group I,A. arguta, A. melanandra, A. kolomikta, and A. marcrosperma, characterized by the non-hair in fruits and loaves or a few pubescences only in young stage, which belongs to the section Leiocarpae, and group II, A. chinensis, A. deliciosa, and A. eriantha, characterized by a lot of hairs only in young fruit stage and with a lot of hairs or fuzzes in leaves and branches, which belongs to the section Stellatae. Group II especially belongs to the series Perfectae of the section Stellatae and was divided into two subgroups; subgroup I containing A. chinensis and A. deliciosa, and subgroup II containing A. eriantha. In contrast, the two species, A. chinensis and A. deliciosa, which are known to have common parents, were divided into two independent subgroups with 80％ of a similarity value. On the other hand, we selected KT6F for variety specific bands, KT12E primers for 'Hayward' and 'Tomuri'. KT7F or KT12F primers were useful for analysis of inheritance pattern in kiwifruit cross-breeding. We suggest that these primers will be a powerful tool for elucidating phylogenetic relationship and selection of novelty kiwifruit in a breeding program.

• Applied Biological Chemistry
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• v.48 no.4
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• pp.335-338
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• 2005

For the development of qualitative PCR detection method of genetically modified rice (Oryza sativa L.), rice species specific gene, SAMDC1 (S-adenosylmethionine decarboxylase), was selected and validated as suitable for use as an endogenous reference gene in rice. The primer pair OsSAMDC1-5'/3' with 110 bp amplicon was used for amplification of the rice endogenous gene, SAMDC1 and no amplified product was observed from 19 different plants as templates. Qualitative PCR method was assayed with 2 different GM rices (Milyang 204 and Iksan 483) developed in Korea. For the qualitative PCRs, the construct-specific detection primer pairs were constructed. Os204-5'/OsNOS-3' amplifying the junction region of GUS gene and NOS terminator introduced in Milyang 204 gave rise to an amplicon 172 bp; also, Os483-5'/OsNOS-3' amplifying the junction region of Bar gene and NOS terminator introduced in Iksan 483 gave rise to an amplicon 161 bp.

• Journal of Microbiology and Biotechnology
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• v.25 no.9
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• pp.1401-1409
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• 2015

The aim of this study was to develop a SYBR Green-based real-time PCR assay for the rapid, specific, and sensitive detection of Acidovorax avenae subsp. citrulli, which causes bacterial fruit blotch (BFB), a serious disease of cucurbit plants. The molecular and serological methods currently available for the detection of this pathogen are insufficiently sensitive and specific. Thus, a novel SYBR Green-based real-time PCR assay targeting the YD-repeat protein gene of A. avenae subsp. citrulli was developed. The specificity of the primer set was evaluated using DNA purified from 6 isolates of A. avenae subsp. citrulli, 7 other Acidovorax species, and 22 of non-targeted strains, including pathogens and non-pathogens. The AC158F/R primer set amplified a single band of the expected size from genomic DNA obtained from the A. avenae subsp. citrulli strains but not from the genomic DNA of other Acidovorax species, including that of other bacterial genera. Using this assay, it was possible to detect at least one genomeequivalents of the cloned amplified target DNA using 5 × 10⁰ fg/µl of purified genomic DNA per reaction or using a calibrated cell suspension, with 6.5 colony-forming units per reaction being employed. In addition, this assay is a highly sensitive and reliable method for identifying and quantifying the target pathogen in infected samples that does not require DNA extraction. Therefore, we suggest that this approach is suitable for the rapid and efficient diagnosis of A. avenae subsp. citrulli contaminations of seed lots and plants.