• 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 Korean Journal of Mycology
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• v.38 no.1
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• pp.34-39
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• 2010

We have attempted to verify 30 strains of Hypsizigus marmoreus from various mushroom stocks in Korea using random amplified polymorphic DNA (RAPD) methodology. Chromosomal DNAs of them were extracted and subjected to PCR analyses with 3 random primers. Each PCR produced approximately 30 distinct PCR bands with the size from 200 bp to 3000 bp. A dendrogram was acquired using the unweighted pair-group method with arithmetic average (UPGMA) clustering methodology on the basis of the DNA band pattern. The analysis revealed that 30 strains of H. marmoreus were clustered into two distinct clusters. Cluster 1 contained 3 subgroups while the cluster 2 consisted of rather diverse strains. Interestingly, Hm3-10, a wild strain collected from Deog-Yu mountain, was not included in either clusters, indicative of uniqueness of this strain. We nextly attempted to develop strain-specific DNA markers to verify a specific strain. A unique band in the RAPD gel lane of Hm0-4 was extracted and its sequence was determined. PCR with a primer set from the determined sequence revealed that the primer set gave a 250 bp DNA band only for Hm0-4, indicating that this approach works well for the strain-specific identification of H. marmoreus.

• Korean Journal of Poultry Science
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• v.20 no.4
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• pp.177-188
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• 1993

This study was performed to find out the reasonable sexing methods In the chicken, obtain the basic information for the mechanisms related to chicken sexual differentiation and identify the genes which known to involved in chicken sex differentiation. The chromosome analysis of chicken embryonic fibroblast was a simple method to determine sex of chicken by means of Z and W chromosome identification. The bands of female chicken genomic DNA digested with Xho Ⅰ and Eco RI restriction endonuclease showed to be useful in direct sex determination and these repetitive sequences of Xho Ⅰ and Eco RI families were proposed to be very homologous in their sequences by colony hybridization analysis. Seven of 150 random primers were selected to amplify the W chromosome-specific band by using arbitrary primed PCR and three of them were useful to identify the sex of chicken. To identify the sex differentiation genes in the chicken, PCR for the amplification of ZFY and SRY sequences was performed. ZFY and SRY sequences were amplified successfully in the chicken genome, implying that chicken genome might have the sex-related conserved sequences similar to mammalian ones. The PCR products of ZFY amplification were the same in both sexes, suggesting that these sequences may be located on autosome or Z chromosome. The profile of PCR amplification for SRY sequences showed variation between sexes, but this result was not enough to specify whether the SRY gene in chicken is on the autosome or sex chromosome.

• Food Science of Animal Resources
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• v.29 no.5
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• pp.647-652
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• 2009

A duplex PCR technique was applied for specific identification of cow and goat milk in milk products by using primers targeting the mitochondrial 12S rRNA gene. Duplex PCR using primers specific for cow and goat generated specific fragments of 223bp and 326bp from cow and goat milk DNA, respectively. Duplex PCR was applied to 15 milk products purchased from the market to verify label statements. The labeling statements of four market milk products, three yoghurt products, and one whole milk powder product were confirmed in the duplex PCR. The labeling statements of five of seven infant milk powder products were also confirmed by duplex PCR but the other two products were shown to be contaminated with either cow or goat milk. The proposed duplex PCR provides a rapid and sensitive approach to detection of as little as 0.1% cow milk in goat milk and one-step detection of cow or goat milk in milk products.

• The Korean Journal of Mycology
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• v.46 no.4
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• pp.467-477
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• 2018

Colletotrichum circinans, an anthracnose pathogen, causes serious damage to onions worldwide. In this study, specific molecular markers were developed to detect C. circinans accurately and quickly with both conventional and real-time PCR methods. The cirTef-F/cirTef-R and cirTu-F/cirTu-R primer sets, which are specific for C. circinans, were constructed by analyzing $tef-1{\alpha}$ and ${\beta}-tubulin$ genes in the fungus. Using the conventional PCR method, 100 pg and 1 ng of fungal DNA could be detected using the cirTef-F/cirTef-R and cirTu-F/cirTu-R sets, respectively. Using the real-time PCR method, 10 pg and 100 pg of fungal DNA could be detected more sensitively with the cirTef-F/cirTef-R and cirTu-F/cirTu-R sets, respectively. Detection of C. circinans from the artificially infected onion seeds was possible by using both conventional and real-time PCR methods and the developed cirTef-F/cirTef-R primer set. The PCR markers specific for C. circinans developed in this study may enhance the efficiency of fungal pathogen detection in imported vegetables and seeds.

• Journal of Korean Society of Forest Science
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• v.96 no.4
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• pp.425-431
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• 2007

To investigate genetic diversity and PCR detection of Rhizina undulata, PCR detection and sequence analysis of rDNA ITS region of R. undulata in soil were analyzed and developed. The length of partial 18S rDNA from four R. undulata isolates were 1,375 nt. The sequence similarity of R. undulata isolates was 100%. The rDNA ITS regions of R. undulata isolates were 585 nt long. Nucleotide sequencing of the ITS regions showed that PDK-1, PTT-1 and PDJ-9 isolates had 100% sequence identity. But, PDS-5 isolate differed from the three isolates by two nucleotide substitution. R. undulata-specific primers designed by the sequence of ITS region were used in PCR detection of R. undulata. PCR products about 525 bp size, which is specific to R. undulata, were amplified from total DNAs of R. undulata isolates. To assay the sensitivity of PCR detection by R. undulata ITS-specific primer, purely cultured mycelial suspension of R. undulata was serially diluted and mixed with 100g of sterile sandy loam soil, respectively. And then, PCR products of total DNAs extracted from each mycelium-soil mixtures were analysed. The PCR protocol could detected up to 1ng mycelium of R. undulata within 100g of soil.

• The Plant Pathology Journal
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• v.19 no.2
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• pp.106-110
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• 2003

Stolbur Phytoplasma was detected from Lilium Oriental hybrids showing flattened stem and flower clustering. The presence of phytoplasma was demonstrated using polymerase chain reaction（PCR) assays with phyto-plasma-universal（P1/P6）and stolbur phytoplasma-specific 16F1/R1-S primer pairs amplifying phytoplasma 16S rDNA regions. Nucleotide suquences of the phytoplasma 16S rDNA were determined. Nucleic acid extracted from lily amplified 1.5 kb DNA with a phytoplasma universal primer pair. In nested PCR, 1.1 kb PCR product was obtained using specific primer pair, indicating an isolate of stolbur phytoplasma. Nucleotide sequence of phytoplasma 16S rDNA reported in this study showed 99.5% and 99.1％ identities with two known stolbur phytoplamas (16Sr XII-A). Also, it exhibited a sequence homology of 98.0％ with phormium yellow leaf (16Sr XII-B), and 97.9％ with Australian grapevine yellows (16Sr XII-B). Meanwhile, it showed 98.1% identity with strawberry green petal phytoplama, (16Sr1-C), and 94.7 % with American aster yellows (16Sr1-B). Homology percentage of the 16S rDNA nucleotide sequence suggests that this phytoplama could be classified into the stolbur phytoplasma, subgroup A (16Sr XII-A), as a type strain stolbur.

• Research in Plant Disease
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• v.18 no.2
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• pp.129-132
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• 2012

Citrus bacterial canker is an economically important disease affecting citrus production in many citrusgrowing areas and several pathotypes have been recognized within the Xanthomonas pathogens causing canker. In view of the containment of the disease, accurate identification of the causal bacterium is important. In this study, triplex PCR method was developed by using the previously reported primers. Two groups of primer combination, such as, one group including primers 2/3, J-pth1/J-pth2 and XACF/XACR, and another group 2/3, J-pth1/J-pth2 and Xac01/Xac02, were suitable for the detection and differentiation of X. a. pv. citri $A^w$, X. a. pv. aurantifolii B and C, and X. a. pv. citrumelo E strains. Moreover, the primer combination of Xac01 and J-pth2 promised us to use as a specific primer set to detect X. a. pv. citrumelo E strain. The PCR methods developed in this study could be used for the rapid differentiation of Xanthomonas pathotypes of citrus.

• The Plant Pathology Journal
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• v.15 no.5
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• pp.287-294
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• 1999

A technique based on the polymerase chain reaction (PCR) for the specific detection of genus Phytophthora and Phytophthora cryptogea-P. drechsleri complex group was developed using nucleotide sequence information of ribosomal DNA (rDNA) regions. The internal transcribed spacers (ITS) including 5.8S were sequenced for P. cryptogea-P. drechsleri complex group and its related species. Two pairs of oligonucleotide primers were designed. Primer pair ITS1/Phy amplified ca. 240 bp fragment in 12 out of 13 specie of Phytophthora, but not in Pythium spp., Fusarium spp.and Rhizoctonia solani. Primer pair rPhy/Pcd amplified 549 bp fragment only in P. cryptogea-P. drechsleri complex group, but not in other Phytophthora spp.and other genera. Specific PCR amplification using the primers was successful in detecting Phytophthora and P. cryptogea-P. drechsleri complex group in diseased plants.

• Applied Biological Chemistry
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• v.50 no.4
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• pp.276-280
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• 2007

Buckwheat often causes severe allergic reactions in sensitive people. One of the major allergenic proteins in common buckwheat (Fagopyrum esculentum) has been found to be a BW10KD protein. In this study, we developed a PCR method to detect buckwheat ingredients in food using primers corresponding to the allergenic BW10KD gene. Five pairs of oligonucleotide primers successfully enabled PCR amplification of the specific regions of the genomic BW10KD DNA from buckwheat, but no amplification from seven other cereals and beans (barley, wheat, German millet, African millet, soybean, red bean, and black bean). The proposed PCR method was applied to analyze 12 processed foods (buckwheat flour, buckwheat noodle, buckwheat jelly, wheat noodle, instant noodle, black sesame gruels, sunsik, cookie, misutkaru, and three kinds of cereal); among them, only three samples including buckwheat flour, buckwheat noodle and buckwheat jelly showed a positive reaction to the detection. This PCR method was able to detect as little as 1 ng of common buckwheat DNA. This rapid and specific PCR method would be applicable to detect allergenic buckwheat ingredients in food.