• The Plant Pathology Journal
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• v.20 no.4
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• pp.302-307
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• 2004

Three pairs of specific primers were developed for rapid and precise RT-PCR detection of three seed-transmissible viruses, namely Peanut clump virus (PCV, Pecluvirus), White clover mosaic virus (WCIMV, Potexvirus) and Carrot red leaf virus (CaRLV, Luteovirus). Each primer set was found in conserved region through multiple sequence alignment in the DNAMAN. Total nucleic acids extracted from PCV-, WCMV-, and CaRLV-infected seeds and healthy plants were used for RT-PCR detection using each virus-specific primer, Sizes of PCV, WCIMV, and CaRLV PCR products were 617bp (PCV-uni5 and PCV-uni3 primers), 561bp (WCMV-CP5 and WCMV-CP3 primers), and 626bp (CL1-UP and CL2-DN primers); which corresponded to the target sizes. Nucleotides sequences of each amplified cDNA were confirmed which belonged to the original virus. This study suggests that these virus-specific primer sets can specifically amplify viral sequences in infected seeds. Thus, they can be used for specific detection of three viruses (PCV, WCMV and CaRLV) from imported seed samples for plant quarantine service.

• Mycobiology
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• v.31 no.4
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• pp.196-199
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• 2003

A very reliable and specific method for the identification of fungi in ectotrophic mycorrhizal symbiosis was developed using a specific PCR assay based on the amplification of the ITS1 region. To obtain specific data, an ITS-diagnostic assay was carried out that reveals genera and species specific sequences. Here, an application of one method is presented, which covers the identification of pure mycelia, basidiocarps as well as mixed samples such as ectomycorrhizal roots that were mingled with remains of the host plant. For this purpose a protocol was established that allowed the extraction of DNA from single mycorrhizal roots. In order to perform a specific ITS analysis we generated a new ITS-primer(ITS8) by a multiple alignment of five different genera and species of mycorrhizal fungi. The utilization of ITS1 and ITS8 resulted in specific PCR amplicons, which were characterized by sequencing without purification steps, even when the template DNA was associated with roots.

• The Korean Journal of Mycology
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• v.44 no.4
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• pp.365-370
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• 2016

We collected leaves of Pinus koraiensis and Thuja koraiensis and roots of Bletilla striata from various sites in Korea. The leaf and root samples were surface-sterilized and endophytic fungi were isolated. Fungal isolates were identified based on their morphological characteristics and a phylogenetic analysis of internal transcribed spacer regions, large subunit regions, and the ${\beta}$-tubulin gene. Consequently, we identified five species of endophytic fungi, namely Colletotrichum simmondsii, Fusarium sterilihyphosum, Diatrypella pulvinata, Ochroconis globalis, and Sphaeria chrysosperma. These species have not been previously reported in Korea and we report them here with descriptions and illustrations.

• Mycobiology
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• v.34 no.1
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• pp.7-13
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• 2006

Arbuscular mycorrhizas (AM) have mutualistic symbiosis with plants and thus efforts have been placed on application of these symbiotic relationships to agricultural and environmental fields. In this study, AM fungi were collected from 25 sites growing with 16 host plant species in Korea and cultured with Sorghum bicolor in greenhouse condition. AM fungal spores were extracted and identified using both morphological and molecular methods. Using morphological characters, total 15 morpho-speices were identified. DNA was extracted from single spore of AM fungi and a partial region on 18S rDNA was amplified using nested PCR with AM fungal specific primers AML1/AML2. A total of 36 18S rDNA sequences were analyzed for phylogenetic analysis and 15 groups of AM fungi were identified using both morphological and molecular data of spores. Among the species, 4 species, Archaeospora leptoticha, Scutellospora castanea, S. cerradensis, S. weresubiae were described for the first time in Korea and two species in Glomus and a species in Gigaspora were not identified. Morphological and molecular identification of AM fungal spores in this study would help identify AM fungal community colonizing roots.

• Korean Journal of Environmental Biology
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• v.21 no.1
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• pp.77-85
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• 2003

We have examined the 16S-23S rRNA intergenic spacer regions (ISR) of Vibrio fluvialis. ISRs were PCR amplified, cloned into a plasmid vector and then sequenced. As results of ISR nucleotide sequence analysis, total of 6 clones were isolated depending on the size. The clones were different in both the number and the composition of the tRNA genes, and were designated ISR-A, ISR-E, ISR-El, ISR-lA, ISR-EKV, ISR-EKAV. ISR-A contains $tRNA^{Ala}$; ISR-lA, $tRNA^{Ile}$-$tRNA^{Ala}$; ISR-EKV, $tRNA^{GIu}$-$tRNA^{Lys}$-$tRNA^{Val}$;ISE-EKAV, $tRNA^{GIu}$-$tRNA^{Lys}$-$tRNA^{Ala}$-$tRNA^{Val}$; ISR -E and E1, $tRNA^{GIu}$ clusters. ISR-EKV was shown to be a minor type out of the six ISR types and showed a very limited homology between ISR-EKV from V, fluvialis and ISRa from other Vibrio species. Therefore ISR-EKV sequence was used to design species-specific primers to detect V, fiuvialis from other Vibrio species by PCR reaction. The specificity of the primers was examined using genomic DNA of other Vibrios as templates for PCR reaction. The result showed that PCR can be a useful method to detect V. fluvialis among Vibrio species in a single PCR reaction.

• Korean Journal of Microbiology
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• v.36 no.3
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• pp.173-180
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• 2000

The intern1 spacer regions (ISR) between the 16s and 23s $1_RNA$ genes of Aeronzonus iwonii blogroupsobria and A. caviae were investigated by PCR fragment length typing and DNA sequencing. A. iwonii bv.sobria has a speciIic 16s-23s pattern of 2-4 fiagments ranging Goin 479-539 bp, with the exception of thespecies Aeron7onns cmiae, which has 3 fragments ranglog from 470-602 bp. In all of the.4 vei*onii bv. sobr,iaand A, caviae strains examined in this study, the 470-481bp Tragnent, designated TSR-1, invariably contained $tDNA^{uc(GAT)$ and $tDNA^{Ala(TGC)$ in contrast to ISR-2 (513-525 bp). ISR-3 (537-539 bp) and ISR-4 (568-602 bp)containing TEX>$tDNA^{Olu(ITC)$ A stretch of 20 nucleotides (178-197 bp) in the ISR-4 was conserved only wit11mA.caiiue, from which the A. caiiae specific primer, named prAC-F, was designed and used for PCR with aAcaviae coimnon reverse primer A PCR product of 450 bp was apparent alnong I , caiizne strains, but not ii1.4.ijeronii bv. sob~ia strains. The PCR product was oot detected t"-om strains belonging to A. hjili-o~~hila, Ebrio,aud the family Ef\ulcornertei,obncteriaceae. This study provides the first molecular tool for mdentifying the species 8.caviae.ing the species 8. caviae.

• Journal of Microbiology and Biotechnology
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• v.17 no.3
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• pp.468-473
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• 2007

White rot fungi, Phanerochaete chrysosporium and Phanerochaete sordida, have been mostly studied in a variety of industrial processes like biopulping and pulp bleaching as well as in bioremediation. Whereas P. sordida is widely distributed in the North Temperate Zone, P. chrysosporium is reported in the restricted area and hundreds of reports have been described from a few strains of P. chrysosporium, which are deposited at various fungal collections in the world. The isolates of two species are not easily discriminated because of their morphological and molecular similarity. Through the ITS sequence analyses, a region containing substantial genetic variation between the two species was identified. PCR amplification using two specific primers was successfully used to differentiate P. chrysosporium from P. sordida. These results were supported by cultural studies. The growth rates at $37^{\circ}C$ on PDA, MEA, and Cza and the microscopic features of conidia on PDA and YMA were also very useful to differentiate those two species.

• Journal of Crop Science and Biotechnology
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• v.10 no.1
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• pp.27-32
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• 2007

RAPD, Inter-SSR, and AFLP markers were used to assess the genetic diversity of lettuce cultivars and the phylogenetic relationships in Lactuca spp. A total of 216 polymorphic bands from seven RAPD primers, four Inter-SSR primers, and five AFLP primer combinations were used to elucidate the genetic similarity among lettuce cultivars. Forty-four lettuce accessions were subdivided into discrete branches according to plant type: crisphead, butterhead, and stem type, with some exceptions. The leafy- and cos-type accessions were intermingled in other groups with no discrete branch indicating that these are more diverse than others. Three accessions, including the Korean cultivar 'Cheongchima', the Korean local landrace 'Jinjam', and the German cultivar 'Lolla Rossa' were classified as the most diverse accessions. Twenty bands were unique in specific cultivars. Among these, three were specific in a plant type; one in Korean leafy type, one in crisphead type, and one in cos type lettuce. In the phylogenetic analysis among Lactuca species, L. saligna, L. serriola, and L. georgica clustered in a sister branch of the L. sativa complex. Two L. virosa accessions show the highest intra-specific relationships. L. perennis outlied from all the other Lactuca species at a genetic similarity of 0.53 and clustered with two Cichorium species, C. intybus and C. endivia, with genetic similarity of 0.67. The phylogenetic tree was supported by data from polymorphism of chloroplast genome which was revealed by PCR-RFLP.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 2003.10a
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• pp.134.2-135
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• 2003

The plant-pathogenic species S. scabies, S. acidiscabies, and S. turgidiscabies cause the scab disease of potato and produce the phytotoxins, thaxtomins. necl, a gene conferring a necrogenic phenotype, is involved in pathogenicity and physically linked to the thaxtomin A biosynthetic genes. Identification of the pathogenic strains of Streptomyces from soil was performed through the polymerase chain reaction by using specific pathogenicity primer sets derived from the necl gene sequences of Streptomyces smbies. The DNA was extracted from soil using a bead-beating machine and modifications of the FastPrep system. The DNA was suitable for direct use in the PCR. The PCR products showed the bands of approximately 460 bp. This methods can be very usuful in identifying species responsible for scab diseases and studying on the ecology of plant-pathogenic Streptomyces spp.

• Journal of Microbiology and Biotechnology
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• v.18 no.2
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• pp.194-198
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• 2008

A newly designed Cytophaga-Flavobacteria-specific 16S rRNA gene primer pair was employed to investigate the CF community structure in the Bering Sea, revealing a previously unknown and unexpected high CF diversity in this high latitude cold sea. In total, 56 clones were sequenced and 50 unique CF 16 rRNA gene fragments were obtained, clustering into 16 CF subgroups, including nine cosmopolitan subgroups, five psychrophilic subgroups, and two putatively autochthonous subgroups. The majority of sequences (82%) were closely related to uncultured CF species and could not be classified into known CF genera, indicating the presence of a large number of so-far uncultivated CF species in the Bering Sea.