• Title/Summary/Keyword: multiplex-PCR typing

Search Result 29, Processing Time 0.033 seconds

Genetic analysis of Campylobacter jejuni isolates from diarrhea patients in Gyeonggi-do (경기도에서 분리된 Campylobacter jejuni의 유전자 패턴 분석 연구)

  • Kim, Woon-Ho;Choi, Ok-Kyung;Jeong, Jin-A;Park, Sung-Hee;Lee, Yea-Eun;Park, Gwang-Hee;Yoon, Mi-Hye
    • Korean Journal of Microbiology
    • /
    • v.54 no.1
    • /
    • pp.31-37
    • /
    • 2018
  • Campylobacter jejuni is an important food-borne pathogen causing gastroenteritis in human. We isolated 208 strains of Campylobacter jejuni from 430 diarrhea patients and food employees with 17 food-poisoning outbreaks between 2014 and 2016 in Gyeonggi area. The strains were tested for genetic relationship and the genotype distribution using PFGE and multiplex-PCR typing. Among the 47 Penner-serotypes known for C. jejuni, it was identified as a genotype consisting of 35 genotypes by multiplex-PCR typing and represented 7 genotypes (HS2, HS4A, HS8, HS15, HS29, HS41, and HS53) in the selected strain. From the PFGE analysis of 11 food-poisoning outbreaks, 5 group of PFGE profile were obtained, and genetic similarity in these clusters ranged from 61.8 to 66.6%. This study examines the genetic diversity of C. jejuni that have been separated in the Gyeonggi area through various genetic analysis methods and identifies the correlation between strains in patients who have been infected with the disease in the future.

Personal identification of the excavated ancient human bone through molecular-biological methods (분자생물학적 방법을 통한 출토인골의 개인 동정-사천 늑도 출토 인골과 민통선 민묘 출토 인골을 중심으로)

  • Seo, Min-Seok;Lee, Kyu-Shik;Chung, Yong-Jae;Lee, Myeong-Hui
    • 보존과학연구
    • /
    • s.22
    • /
    • pp.27-40
    • /
    • 2001
  • DNA typing is often used to determine identity from human remains. Recently, the molecular biological analysis of ancient deposits has become possible since methods for the recovery of DNA conserved in bones or teeth from archaeological remains have been developed. In the field of archaeology, one of the most promising approaches is to identify the individuals present in a mass burial site. We performed nuclear DNA typing and mitochondrial DNA sequencing analysis based on PCR from a Korea ancient human remain excavated from Sa-chon Nuk-island and civilian access controlline(CACL). A femur bone were collected and successfully subjected to DNA extraction, quantification, PCR amplification, and subsequently typed for several shot tandem repeat(STR)loci. 4 types of STR systems used in this study were CTT multiplex(CSF1PO, TPOX, TH01), FFv multiplex(F13A01, FESFPS, vWA), Silver STRⅢ multiplex(D16S539, D7S820, D13S317), and amelogenin for sex determination. This studies are primarily concerned with the extraction, amplification, and DNA typing of ancient human bone DNA samples. Also, it is suggestive of importance about closely relationship between both fields of archaeology and molecular biology.

  • PDF

Molecular typing and detection of enterotoxin by multiplex PCR of Staphylococcus aureus isolated from bovine mastitis (유방염 유즙에서 분리한 포도구균의 분자생물학적 typing과 multiplex PCR을 이용한 장독소의 검출)

  • Kim, Sin;Hong, Hyon-Pyo;Kim, Sang-Yun;Kwon, Heon-Il;Lee, Hee-Moo
    • Korean Journal of Veterinary Service
    • /
    • v.25 no.3
    • /
    • pp.275-283
    • /
    • 2002
  • Forty strains of Staphylococcus aureus were isolated from mastitic milk. As a result of antimicrobial susceptibility test, the strains of S aureus revealed 47.5% were resistant to ampicillin and penicillin, and 7.5% to gentamicin. But 45% of isolates were sensitive to antimicrobial agents tested. In case of enterotoxin production, 56.3% of 16 strains produced enterotoxin D. Two strain of enterotoxin D producers produced both enterotoxin B and D. According to isolation date, 15 representative strains were selected. As a results of pulsed field gel eletrophoresis analysis of the 15 representative strains, 14 strains were identical. Therefore we consider the identical strains of S aureus have caused continuously bovine mastitis in this dairy farm. If autogenous vaccine can be made by the strains, it will work well for the prevention of bovine mastitis caused by S aureus.

Simultaneous diagnosis and differentiation of Mycoplasma hyopneumoniae and Mycoplasma hyorhinis infections by multiplex PCR (Mycoplasma hyopneumoniae와 Mycoplasma hyorhinis 동시 감별진단을 위한 다중진단 중합효소반응)

  • Hong, Sunhwa;Lee, Hyun-A;Kim, Dong-Woo;Kim, Tae-Wan;Kim, Okjin
    • Korean Journal of Veterinary Service
    • /
    • v.37 no.4
    • /
    • pp.247-252
    • /
    • 2014
  • The economic impact of swine mycoplasma infection is high. An accurate diagnosis is often difficult and time consuming. We report the development and validation of an effective multiplex polymerase chain reaction (PCR) assay that detects Mycoplasma (M.) hyopneumoniae and M. hyorhinis. The multi detection of M. hyopneumoniae and M. hyorhinis primer set were employed to detect mycoplasma species and typing of the species was performed on the basis of sequence analysis of the PCR product. The target nucleic acid fragments were specifically amplified by M. hyopneumoniae and M. hyorhinis PCR with 16S ribosomal DNA primers. Single and mixed Mycoplasma species DNA templates were used to evaluate the specificity of the multiplex assay. The corresponding specific DNA products were amplified for each pathogen. The multiplex PCR assay provides a novel tool for simultaneous detection and differentiation of M. hyopneumoniae and M. hyorhinis.

Development of Ultra-rapid Multiplex Real-time PCR for the Detection of Genes from Avian Influenza Virus subtype H5N1 (조류인플루엔자 H5N1 바이러스 유전자의 신속 검출을 위한 초고속 다중 실시간 PCR법의 개발)

  • Kim, Eul-Hwan;Lee, Dong-Woo;Han, Sang-Hoon;Lim, Yoon-Kyu;Yoon, Byoung-Su
    • Korean Journal of Veterinary Research
    • /
    • v.47 no.4
    • /
    • pp.399-407
    • /
    • 2007
  • Cause of high lethality and dissemination to human being, new development of rapid method for the detection of highly pathogenic Avian Influenza Virus (AIV) is still necessary. For the detection of AIV subtype H5N1, typical pathogenic AIV, new method to confirm sub-typing of this virus is also needed. For the purpose of ultra-rapid detection and sub-typing of hemagglutinin and neuraminidase of AIV, this study was planned. As the results we could demonstrate an ultra-rapid multiplex real-time PCR (URMRT PCR) for the detection of AIV In this study, the URMRT PCR were optimized with synthesized AIV H5- and AIV Nl-specific DNA templates and GenSpector TMC, which is a semiconductor process technology based real-time PCR system with high frequencies of temperature monitoring. Under eight minutes, the amplifications of two AIV subtype-specific PCR products were successfully and independently detected by 30 cycled ultra-rapid PCR, including melting point analysis, from $1{\times}10^3$ copies of mixed template DNA. The URMRT PCR for the detection of AIV H5N 1 developed in this study could be expected to apply not only detections of different AIVs, but also various pathogens. It was also discussed that this kind of the fastest PCR based detection method could be improved by advance of related technology in near future.

Detection and Typing of Human Papillomavirus in Cutaneous Common Warts by Multiplex Polymerase Chain Reaction (Multiplex PCR 기법을 이용한 보통사마귀 내 인유두종바이러스 검출 및 분류)

  • Choi, Soon-Yong;Lim, Jong-Ho;Kim, Eun-Jung;Kim, Hei-Sung;Kim, Beom-Joon;Kang, Hoon;Park, Young-Min
    • Journal of Life Science
    • /
    • v.21 no.7
    • /
    • pp.947-952
    • /
    • 2011
  • A number of epidemiological studies have identified human papillomavirus (HPV) types 1, 2, 3, 4, 7, 10, 27, 57, and 65 in cutaneous common warts. However, identification of the HPV subtype by conventional polymerase chain reaction (PCR) is time consuming with its multi-step laboratory process. In this study, we aim to develop a specific one-step multiplex polymerase chain reaction method which capably identifies six different HPV genotypes related to common warts. By HPV DNA sequence analysis, 6 pairs of specific primers were designed from the intergenic regions of genes L1 to E6, and from genes E2 to L2. DNA sequence analysis with the L1 gene sequence of the sample was performed to measure the specificity of multiplex PCR. HPV-1, -2, -3, -4, -27, and -57 were identified without cross amplification in 109 out of 129 samples. The sensitivity and specificity of our set of primers in detecting HPV were 85% and 99.5%, respectively. For the 20 samples where HPV type was not identifiable by our batch of primer sets, multiplex PCR with an additional set of HPV primers was done, where 7 were found positive for HPV-7 or -65. Our results demonstrate that the newly designed multiplex PCR can rapidly detect the specific HPV subtype involved in common warts with high accuracy.

Direct Multiplex Reverse Transcription-Nested PCR Detection of Influenza Viruses Without RNA Purification

  • Song, Man-Ki;Chang, Jun;Hong, Yeong-Jin;Hong, Sung-Hoi;Kim, Suhng-Wook
    • Journal of Microbiology and Biotechnology
    • /
    • v.19 no.11
    • /
    • pp.1470-1474
    • /
    • 2009
  • This paper describes the development a of direct multiplex reverse transcription-nested polymerase chain reaction (PCR) method, devised for simultaneous detection and typing of influenza viruses. This method combines the direct reverse transcription reaction without RNA purification with the enhancement of sensitivity and specificity of nested PCR. The method successfully detected three major human influenza viruses: influenza virus A subtype 1 (H1N1) and subtype 3 (H3N2), and influenza B virus (B). The minimum number of virus particles (pfu/ml) necessary for detection in spiked saliva samples was 200 (H1N1), 140 (H3N2), and 4.5 (B). The method's sensitivity and simplicity will be convenient for use in clinical laboratories for the detection and subtyping of influenza and possibly other RNA viruses.

Molecular Biological Characteristics of Vibrio cholerae O1 Isolated from Diarrheal patients in the Gyeongbuk province. (최근 경북지역 설사환자 검체에서 분리된 Vibrio cholerae O1의 분자생물학적 특성)

  • 이상조;이복권;이건주;이희무
    • Microbiology and Biotechnology Letters
    • /
    • v.31 no.4
    • /
    • pp.334-341
    • /
    • 2003
  • This study was carried out to investigate the cause of cholera outbreak in Gyeongbuk province in 2001.90 strains of Vibrio cholerae O1 El Tor serotype Inaba were isolated from diarrheal patients. By multiplex-PCR, all of the isolated strains revealed positive for detection ctxA, hlyA and tcpA genes. There were DNA sequence difference of the cholera-toxin subunit A gene and subunit B gene between isolated V. cholerae O1 and the strain of GenBank. In analysis of PFGE patterns, all of the isolated strains were showed the same DNA fragments. We also collected plankton samples in the east coast of Gyeongbuk to isolate V. cholerae O1 and V. cholerae O139 from August to October 2002. The samples were examined to detect the rfb gene and cholera-toxin gene by multiplex-PCR. The cholera-toxin gene was detected and then we tried to isolate V. cholerae O1 and V. cholerae O139, but they were not isolated.

Gene typing of staphylococcal superantigens produced by Staphylococcus aureus isolates from meat (식육유래의 Staphylococcus aureus으로부터 산생되는 staphylococcal superantigens의 유전자형 분석)

  • Yoon, Jang-won;Jung, Suk-chan;Yang, Soo-jin;Jung, Byong-youel;Seo, Keon-suk;Kim, So-hyun;Park, Yong-ho
    • Korean Journal of Veterinary Research
    • /
    • v.38 no.3
    • /
    • pp.553-558
    • /
    • 1998
  • Using the previous established multiplex PCR (mPCR), the presence of six kinds of staphylococcal superantigen (SAg) genes was investigated for nineteen Staphylococcus aureus isolates from the commercialized meat sources. As a result, only one isolate from pork among 19 S aureus isolates (5.3%) was confirmed as a potential SAg producer and harbored sec gene. The results in this study suggest that meat may not be major contagion of staphylococcal food poisoning (SFP) in Korea and that staphylococcal enterotoxin type C may be associated with the disease. Also, the mPCR method in this study can be a useful genotypic method which can overcome the typical disadvantages of conventional antibody-based methods due to antigenic homology, and furthur survey on food-borne S aureus isolates can provide the important epidemiological data for SFP in Korea.

  • PDF