• Journal of fish pathology
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• v.21 no.3
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• pp.181-188
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• 2008

Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay was used to detect and identify four fish viruses, fish iridovirus, viral hemorrhagic septicaemia virus (VHSV), viral nervous necrosis virus (VNNV), hirame rhabdovirus (HRV). Four viruses were detected by PCR with each specific primers. Identification of iridovirus was achieved by digesting the PCR amplified fragment with a restriction enzyme ApaⅠ. It was possible to distinguish positive from false positive PCR amplicons of VHSV by RFLP of PstⅠ or HindⅢ restriction enzymes. VNNV was identified using RFLP of BamHⅠrestriction enzyme and HRV was identified by XbaⅠ restriction enzyme. This approach can be used for more rapid, simple and specific diagnosis of fish viral diseases.

• Journal of Food Hygiene and Safety
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• v.23 no.4
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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.

• Fisheries and Aquatic Sciences
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• v.11 no.3
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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.

• Journal of Periodontal and Implant Science
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• v.29 no.2
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• pp.349-355
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• 1999

The present study has been performed to develop a PCR technology to identify human immunoglobulin(Ig) allotypes with restriction fragment length polymorphism(RFLP) using a probe. Genomic DNA were ampilified with PCR tecnology using primers from peripheral blood lymphocytes of 10 periodontal patiens, whose Ig allotypes have been pre-determined by serological tecnique using heagglutination technique. The result indicated that the RFLP patterns could successfully differentiate the Ig allotypes, which suggests that this technology can be developed as a tool useful for population genetics studies.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.2
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• pp.262-266
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• 2017

Objective: This study was conducted to screen scrotal hernia in domesticated swine from selected breeders in the Philippines. This defect is associated with a cytosine to thymine mutation in the BCL-2 associated X protein (BAX) gene of swine. Methods: Genetic screening was done by DNA extraction followed by amplification and digestion using polymerase chain reaction-restriction fragment length polymorphism, amplifying the 416 bp region of the BAX gene that was subjected to digestion using the Ear I enzyme. Sequencing was also conducted to validate the results. Results: Results revealed that out of 538 samples tested, 411 (76.4%) of the samples were found to be normal whereas the remaining were carriers of the mutation in which 80 (14.9%) were heterozygous mutants and 47 (8.7%) were homozygous mutants. Pietrain breed was found to have the highest incidence. Conclusion: Having a scrotal hernia eliminates the chances of using the boar as a breeder stock because the following generations arising from it would most likely exhibit herniation. It is therefore advised to establish a genetic screening method for Scrotal Hernia in the Philippines to eliminate the negative gene from the herd.

• Asian-Australasian Journal of Animal Sciences
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• v.27 no.10
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• pp.1487-1492
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• 2014

This research applied and evaluated a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using cytochrome b gene to detect pork contamination in meatballs from local markets in Surabaya and Yogyakarta regions, Indonesia. To confirm the effectiveness and specificity of this fragment, thirty nine DNA samples from different meatball shops were isolated and amplified, and then the PCR amplicon was digested by BseDI restriction enzyme to detect the presence of pork in meatballs. BseDI restriction enzyme was able to cleave porcine cytochrome b gene into two fragments (131 bp and 228 bp). Testing the meatballs from the local market showed that nine of twenty meatball shops in Yogyakarta region were detected to have pork contamination, but there was no pork contamination in meatball shops in Surabaya region. In conclusion, specific PCR amplification of cytochrome b gen and cleaved by BseDI restriction enzymes seems to be a powerful technique for the identification of pork presence in meatball because of its simplicity, specificity and sensitivity. Furthermore, pork contamination intended for commercial products of sausage, nugget, steak and meat burger can be checked. The procedure is also much cheaper than other methods based on PCR, immunodiffusion and other techniques that need expensive equipment.

• Parasites, Hosts and Diseases
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• v.34 no.1
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• pp.15-20
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• 1996

Cysts of Entamoebn histoIMtica are still found from humans in Korea, but notall of the cysts are known as pathogenic. The non-pathogenic strain is regarded as a differenL species, E. nispnr. In this study, Korean isolates of conventional E. histolvticn were subjected for the differentiation by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis. Human stools were screened by routine microscopic examination, and cyst or trophozoite positive stools were inoculated into Robinson media. The cultivated trophozoites were prepared for DNA extraction, and the DNAs were used for PCR with common primers of Pl gene. The PCR products were divested with 3 restriction enzymes and RFLP was observed. Also anti-sense primers containing the cleavage site of each restriction eWe were designed for differentiation only by PCR. The PCR products of Korean isolates 59,512, YS-6, and YS-27 were spliced by Taq I and Xmnl but not byAccl, and the isolates S1, S3, S11, S15, S16, S17, S20, YS- l7, and YS-44 were spliced by Acc I but not by Taq I and Xmn I. These RFLP pattern correlated well with PCR products by the species specific primers. The findings confirm that the Korean isolates 59,512, YS-6, and YS-27 are E. histolwtico and others are E. dispar. In Korea, most of the asymptomatic cyst carriers are infected by E. dispar, not by E. histolytica. Key words: Entcmoebc histolytica, Entcmoebn dispar Korean isolates, polymerase chain reaction (PCR), restriction fragment length polymorphism (RFLP)

• Korean Journal of Clinical Laboratory Science
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• v.47 no.2
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• pp.83-89
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• 2015

Although Mycobacterium tuberculosis complex strains remain responsible for the majority of diseases caused by mycobacterial infections worldwide, the increase in HIV (human immuno deficiency virus) infections has allowed for the emergence of other non-tuberculous mycobacteria as clinically significant pathogens. M. tuberculosis was detected by two-tube nested polymerase chain reaction (PCR) and non-tuberculous mycobacteria was detected by PCR-restriction fragment length polymorphism (RFLP) with Msp I. Result of niacin test is equal to result of two-tube nested PCR after culture for M. tuberculosis. In this study, acid fast bacilli stain (AFB. stain) >2+ case, Detection of Mycobacteria is similar to result before culture and after culture. AFB. stain <1+ case, result of mycobacteria is distinguished. Conclusionly, these results suggest that identification of mycobacteria must go side by side both culture and PCR for more fast and accuracy.

• Restorative Dentistry and Endodontics
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• v.20 no.2
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• pp.577-609
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• 1995

Bacteria have been regarded as one of the most important factors in pulpal and periapical diseases. Streptococci are frequently isolated facultative anaerobes in infected root canals. Recently molecular biological techniques have been rapidly progressed. This study was designed to apply the molecular biological tools to the identification and classification of streptococci in the endodontic microbiology. Streptococci isolated from infected root canals were identified with both Vitek Systems and API 20 STREP. Identification results were somewhat different in several strains of streptococci. Eighteen streptococci and enterococcal was difficult so to digest plasmid DNA using Hind III and EcoRI to differentiate strains by restriction enzyme analysis of plasmid DNA. 16S rDNA of chromosome was amplified by polymerase chain reaction(PCR) and then restricition fragment length polymorphism(RFLP) using several restriction enzymes was observed. The molecular mass of 16S rDNA of chromosomal DNA was approximately 1.4kb. There were three to five RFLP patterns using eight restriction enzymes. RFLP patterns digested with CfoI which recognizes four base sequences were identical in all stains. Hind III which recognizes six base sequences could not digest the 16S rDNA. Restriction enzymes which recognize five base sequences were suitable for RFLP pattern analysis. At least three different restriction enzymes were needed to compare each strains. 16S rDNA PCR-RFLP was simple and rapid to differentiate and classify strains and could be used in the epidemiological study of root canal infections.

• Journal of Korean Society on Water Environment
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• v.22 no.4
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• pp.590-598
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• 2006

In-situ Air sparging (IAS) is a groundwater remediation technique, in which organic contaminants volatilize into air form the saturated to vadose zone. This study was carried out to evaluate the effect of sludge and soil microbial community structure on air sparging of Total Petroleum Hydrocarbons (TPH) contaminated groundwater soils. In the laboratory, diesel (10,000 mg TPH/kg) contaminated saturated soil. The Air was injected in intermittent (Q=1500 mL/min, 10 minute injection and 10 minute idle) modes. For Terminal-Restriction Fragment Length Polymorphism (T-RFLP) analysis of eubacterial communities in sludge of wastewater treatment plants and soil of experiment site, the 16S rDNA was amplified by Polymerase Chain Reaction (PCR) from the sludge and the soil. The obtained 16S rDNA fragments were digested with Msp I and separated by electrophoresis gel. We found various sequence types for experiment with sludge soil samples that were closely related to Agrococcus, Flavobacterium, Thermoanaerobacter, Flexibacter and Shewanella, etc, in the clone library. The results of the present study suggests that T-RFLP method may be applied as a useful tool for the monitoring in the TPH contaminated soil the fate of microorganisms in natural microbial community.