• Journal of Microbiology and Biotechnology
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• 제5권4호
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• pp.181-187
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• 1995

The cells of Vibrio vulnificus can be induced to the viable but nonculturable (VBNC) state by natural environmental parameters. The V. vulnificus cells in the VBNC state can not be recovered by ordinary laboratory techniques. This nonculturability could often hamper development of effective processing strategies to minimize the number of V. vulnificus in seafoods. Even with V. vulnificus cells in a culturable state, the length of time required to identify the bacteria in contaminated food by phenotyphic characterization may prevent appropriate in-time responses by public health agencies to infections of the bacteria. In the present study, we used polymerase chain reaction (PCR) to develop a rapid and direct detection method for V. vulnificus in small octopus (Octopus variabilis) which is consumed as a raw food in Korea. The region targeted was a 704-base pair (bp) portion of the hemolysin gene, vvhA, of V. vulnificus. The primers designed for PCR amplification were specific for all V. vulnificus sp. tested. Several methods were examined to extract total DNA directly from V. vulnificus seeded into the octopus homogenate and the guanidine isothiocyanate (CITC) method appeared to be most effective. From the octopus homogenate seeded by V. vulnificus at an initial level of $10^2$ CFU/ml of the homogenate and then incubated for 12 h, the targeted sequence was successfully amplified by PCR and the 704-bp DNA fragment was observed by gel electrophoresis. The total completion of this assay requires less than one day.

• Asian Pacific Journal of Cancer Prevention
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• 제14권9호
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• pp.4989-4992
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• 2013

Aim: To compare p16INK4a immunocytochemistry with the HPV polymerase chain reaction in predicting high grade cervical squamous intraepithelial lesions. Materials and Methods: This diagnostic case-control study was conducted from January 2010 until December 2010. We obtained 30 samples, classified according to the degree of cervical intraepithelial neoplasia (CIN): 11 samples for CIN 1, 9 samples for CIN 2, and 10 samples for CIN 3. HPV PCR, p16INK4a immunocytochemistry, and histopathological examination were performed on all samples. Statistical analysis was conducted using SPSS 20.0. Results: In predicting CIN 2-3, we found p16INK4a to have similar specificity and positive predictive value as HPV PCR (95%, 97.2% vs 96.7%), but better sensitivity (87.5% vs 72.5%) and negative predictive value (82.1% vs 67.6%). The most prevalent types of high-risk HPV in our study were HPV 33, 35, 58, 52, and 16. Conclusions: p16INK4a has better diagnostic values than HPV PCR and may be incorporated in the triage of ASCUS and LSIL to replace HPV PCR. Genotype distribution of HPV differs in each region, providing a challenge to develop HPV vaccines based on the epidemiology of HPV in that particular region.

• Journal of Microbiology and Biotechnology
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• 제12권5호
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• pp.801-806
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• 2002

A multiplex competitive polymerase chain reaction (PCR) method was developed for the rapid identification and quantification of Leuconostoc mesnteroides and Lactobacillus plantarum populations which are the key microorganisms in kimchi fermentation. The strain-specific primers were designed to selectively amplify the target genes encoding 165 rRNA of L. plantarum and dextransucrase of L. mesenteroides. There was a linear relationship between the band intensity of PCR products and the number of colony forming units of each model organism. The PCR quantification method was compared with a traditional plate-counting method f3r the enumeration of the two lactic acid bacteria in a mixed suspension culture and also applied to a real food system, namely, watery kimchi. The population dynamics of the two model organisms in the mixed culture were reliably predictable by the competitive PCR analysis.

• Fisheries and Aquatic Sciences
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• 제11권3호
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• pp.133-139
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• 2008

Molecular techniques, including restriction fragment length polymorphism(RFLP) and polymerase chain reaction-single strand conformational polymorph isms(PCR-SSCP), were developed to identify salted, dried threadfin(Eleutheronema tetradactylum) and white herring(Ilisha elongata) fish. Using PCR with universal primers, conserved 367-bp fragments of the cytochrome b gene were amplified from fresh fish samples and sequenced. The sequences were then searched for specific restriction sites. The digestion of the PCR products with the endonucleases AvaI, FokI, MboII, and MspI generated RFLP, which was used to identify the commercial products. Similarly, the amplified PCR-SSCP products were developed and the products tested. Overall, similar patterns were found in the majority of the fresh and processed products. Based on the results, both RFLP and PCR-SSCP were useful in determining and validating the authenticity of the fish species used to prepare the commercial salted, dried products. A similar approach can be applied to other species.

• 한국식품위생안전성학회지
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• 제23권4호
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• pp.314-318
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• 2008

This research aimed to compare the detection methods of Anisakis simplex in Sea fish by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and macroscopic inspection. We examined 18 Trichiurus lepturus, 11 Scomber japonicus, and 65 Todarodes pacificus collected from the retail markets in the areas of Uljin, Kyuonggi province and Seoul. As the result of examinations, we found that detection rate of Anisakis simplex by macroscopic observation was 89% in Trichiurus lepturus, 90.9% in Scomber japonicus, 32.3% in Todarodes pacificus. The detection rate of Anisakis simplex by PCR-RFLP was 77.7% in Trichiurus lepturus, 81.8% in Scomber japonicus, 26.1% in Todarodes pacificus. We could conclude that PCR-RFLP method of Anisakis simplex was more specific rather than macroscopic observation.

• 한국축산식품학회지
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• 제37권4호
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• pp.599-605
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• 2017

Korean native honey (KNH) is much more expensive than European honey (EH) in Korea, because KNH is a favored honey which is produced less than EH. Food fraud of KNH has drawn attention of the government office concerned, which is in need of a method to differentiate between KNH and EH which are produced by the Asiatic honeybee, Apis cerana and the European honeybee, Apis mellifera, respectively. A method to discriminate KNH and EH was established by using duplex polymerase chain reaction (PCR) in this study. Immunochromatographic assay (IC) was examined to analyze the duplex PCR product. The DNA sequences of primers for the duplex PCR were determined by comparing cytochrome C oxidase genes of the two honey bee species. Chelex resin method was more efficient in extracting genomic DNA from honey than the other two procedures of commercial kits. The duplex PCR amplifying DNA of 133 bp were more sensitive than that amplifying DNA of 206 bp in detecting EH in the honey mixture of KNH and EH. Agarose gel electrophoresis and IC detected the DNA of 133 bp at the ratios of down to 1% and 5% EH in the honey mixture, respectively and also revealed that several KNH products distributed by internet shopping sites were actually EH. In conclusion, the duplex PCR with subsequent IC could also discriminate between KNH and EH and save time and labor.

• 한국동물위생학회지
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• 제24권1호
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• pp.43-50
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• 2001

Swine proliferative enteritis(SPE) caused by inかsoma intracellularis is a common enteric disaese of grower and finisher pig. Swine affected with SPE show variable clinical signs including diarrhea, weight loss, aberrant growth and death. The characteristic lesion of ileitis at necropsy is marked thickening of the last section of the small intestine. The inner lining of the thickened intestine proliferates almost like a cancer and curved rod bacteria(L intracellularis) are always seen inside the intestinal wall. Infected swine shed the organism in the feces. Isolation and growth of pure L intracellularis in vitro requires a suitable cell culture. This procedure is difficult and not a practical means of diagnosis, thus the polymerase chain reaction(PCR) test of feces can be used to determine whether a pig is shedding the infective organism. A sensitive assay based on amplification of a 319bp DffA fragment of the L intracellularis of Swine proliferative enteritis was attempted for the detection of the organism in the 62 feces of swine. L intracellularis was identified on three herds and detected in 6 fecal samples, representing a infection rate of 9.7%. The PCR was very sensitive and specific on the individual level. The PCR technique could be very useful for the diagnosis of this disease.

• 대한수의학회지
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• 제39권2호
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• pp.338-342
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• 1999

Twenty three strains of Escherichia (E) coli O157 : H7 isolated from Korea, Japan, USA were analyzed by pulsed-field gel electrophoresis (PFGE) of XbaI-digested chromosomal DNA and multiplex polymerase chain reaction. Various PFGE patterns of E. coli O157 : H7 were found on the same farm. Most of the E, coli O157 : H7 strains had shiga-like toxin (slt) II gene only (43.5%) or both slt I and slt II genes(30.4%). eaeA gene was highly conserved in the E. coli O157 : H7. There was no correlation between PFGE and slt gene patterns. The results indicate that various genotypes of E. coli O157 : H7 have spread throughout the country and genomic DNA patterns generated by PFGE are highly specific for different strains and have significant value in epidemiologic investigations of infectious disease outbreaks.

• 원예과학기술지
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• 제18권6호
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• pp.797-801
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• 2000

본 연구는 고추에서 Agrobacterium tumefaciens를 이용한 형질 전환시 형질전환율을 높이기 위하여 A. tumefaciens와의 접종시간과 인위적인 상처처리에 따른 효과를 조사하였던 바 고추 형질전환시 자엽절편체의 재분화는 인위적 상처처리 및 A. tumefaciens 접종시간에 의해 영향을 받았다. 접종시간을 3600초로 한 경우의 재분화는 상처처리에 의해 급격히 감소하였으며 자엽 절편체에 대한 인위적인 상처처리가 20초 또는 120초간의 접종시간처리와 병행됨으로써 형질 전환율을 58-72% 이상으로 높이는 효과를 보였다. 형질전환여부의 확인을 ${\beta}$-glucuronidase 염색법과 npt II primer를 이용한 polymerase chain reaction 분석에 의해 수행 한 바 효과적인 것으로 판단되었다.

• 센서학회지
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• 제14권5호
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• pp.337-342
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• 2005

A silicon-based micro-reactor to amplify small amount of deoxyribonucleic acid (DNA) has been fabricated using micro-electro-mechanical systems (MEMS) technology. Polymerase chain reaction (PCR) of DNA requires a precise and rapid temperature control. A Pt sensor is integrated directly in the chamber for real-time temperature measurement and an infrared lamp is used as external heating source for non-contact and rapid heating. In addition to the real-time temperature sensing, PCR needs a rapid thermocycling for effective PCR. For a fast thermal response, the thermal mass of the reactor chamber is minimized by removal of bulk silicon volume around the reactor using double-side KOH etching. The transparent optical property of silicon in the infrared wavelength range provides an efficient absorption of thermal energy into the reacting sample without being absorbed by silicon reactor chamber. It is confirmed that the fabricated micro-reactor could be heated up in less than 30 sec to the denaturation temperature by the external infrared lamp and cooled down in 30 sec to the annealing temperature by passive cooling.