• Title/Summary/Keyword: PCR-SSP

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Genetic Identification of the Kimchi Strain Using PCR-based PepN and 16S rRNA Gene Sequence (PepN과 16S rRNA Gene Sequence 및 PCR 방법을 이용한 김치 젖산균의 동정)

  • Lee, Myung-Ki;Park, Wan-Soo;Lee, Byong-H.
    • Korean Journal of Food Science and Technology
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    • v.32 no.6
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    • pp.1331-1335
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    • 2000
  • The WL6 strain isolated from Kimchi could not be made scientific name because it was identified as three species, i.e., Leuconostoc mesenternides ssp cremoris, Leu. mesenteroides ssp. dextranicum or Lactobacillus bifermentans when it was tested by API kit or Biolog system methods. The unidentifiable WL6 strain was finally reclassified as Lactobacillus bifermentans by genetic identification using two PCR-based specific sequence primer sets which were originated from homologous pepN and 16S rRNA genes.

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Detection of Mycobacterium avium ssp paratuberculosis in Korean Cattle by the Polymerase Chain Reaction (한우 혈액에서 PCR을 이용한 Mycobacterium avium ssp paratuberculosis의 검출)

  • Kim, Kwang-Hyun;Kwak, Kil-Han;Song, Hee-Jong;Cho, Jeong-Gon
    • Journal of Veterinary Clinics
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    • v.27 no.1
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    • pp.23-28
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    • 2010
  • Mycobacterium avium ssp paratuberculosis, intracellular bacteria that can cause chronic granulomatous enteritis in cattle, continues to pose significant economic losses and health problem with high prevalence. The purpose of this study is the polymerase chain reaction (PCR)-base strategy for early detection of M. avium ssp paratuberculosis in whole blood. Blood samples were collected from korean cattles in Jeonbuk, Korea. The 16 out of 88 serum samples were detected M. partuberculosis by ELISA. Then samples of infected 8 Korean cattles were amplified by PCR. The PCR amplified targets are 16s rDNA and heat shock protein 65kDa (hsp 65). The 16s rDNA provided a highly sensitive and specific tool for the direct detection of mycobacteria. In addition M. avium was confirmed characteristically by the hsp65. Finally there were sure to M. avium ssp paratuberculosis by IS900 PCR. The restriction fragment length polymorphism was identified by PCR amplifications and subsequence restriction enzyme digestions with Pst I of a hsp65. These results indicate that confirm M. avium with 16s rDNA, hsp65 and a restriction fragment length polymorphism in the hsp65 gene can be seem the other pattern. Therefore, these results can be used for clinical direct detections of M. avium ssp paratuberculosis in whole blood of Korean cattle and also to be used epidemiological researches.

Genotyping of HLA-DRB1 by Polymerase Chain Reaction-Sequence Specific Primer (Polymerase Chain Reaction-Sequence Specific Primer를 이용한 HLA-DRB1 유전자의 DNA 다형성)

  • Jang, Soon-Mo
    • Korean Journal of Clinical Laboratory Science
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    • v.37 no.3
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    • pp.139-142
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    • 2005
  • Most expressed HLA(human leukocyte antigen) loci exhibit a remarkable degree of allelic polymorphism, which is derived from sequenceing differences predominantly localized to discrete hypervariable regions of the amino-terminal domain of the molecule. In this study, the HLA-DRB1 genotypes were determined in twenty students using the PCR-SSP (polymerase chain reaction-sequence specific primer) technique. Two specific primer pairs in assigning the DRB1 gene were used. The results of PCR-SSP, the $HLA-DRB1^{\ast}0101$ primer detected nine and $HLA-DRB1^{\ast}1501$ primer detected three people. This study shows that the PCR-SSP technique is relatively simple, fast and a practical tool for the determination of the HLA-DRB1 genotypes. Moreover, these genotype frequency results of the HLA DRB1 gene could be useful for database study before being applied to individual identification and transplantation immunity.

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Genotyping of HLA-B by Polymerase Chain Reaction-Sequence Specific Primer (Polymerase Chain Reaction-Sequence Specific Primer를 이용한 HLA-B 유전자의 DNA 다형성 조사)

  • Jang, Soon-Mo
    • Korean Journal of Clinical Laboratory Science
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    • v.39 no.3
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    • pp.147-150
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    • 2007
  • Most expressed HLA (human leukocyte antigen) loci exhibit a remarkable degree of allelic polymorphism, which derives from sequence differences predominantly localized to discrete hypervariable regions of the amino terminal domain of the molecule. In this study, the HLA-B genotypes were determined in twenty students unrelated koreans using the PCR-SSP (polymerase chain reaction-sequence specific primer) technique. Several specific primer pairs in assigning the HLA-B gene were used ($B^{\ast}4001/4007$, $B^{\ast}4901/5001/4501$, $B^{\ast}3701$, $B^{\ast}5801$). The results of PCR-SSP, the HLA-B3701 primer was detected one (5%), the $HLA-B^{\ast}5801$ were detected four (20%), the $HLA-B^{\ast}4001/4007$ were detected nineteen (95%) and the $HLA-B^{\ast}4901/5001/4501$ were detected twenty. This study shows that the PCR-SSP technique is relatively simple, fast and a practical tool for the determination of the HLA-B genotypes. Moreover, these results genotype frequency of the HLA-B gene could be useful for database study before being applied to individual identification and transplantation immunity.

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Molecular Analysis of HLA-C Using Polymerase Chain Reaction-Sequence Specific Primers

  • Lee, Kyung-Ok;Hong, Sung-Hoi;Kim, Min-Jung;Park, Taek-Kyu;Kim, Yoon-Jung;Lee, Kyu-Pum
    • BMB Reports
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    • v.30 no.1
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    • pp.26-32
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    • 1997
  • Of all HLA class I molecules, HLA-C gene products are most poorly understood because they express at a low level on the cell surface compared to HLA-A and -B. In order to identify serologically detectable and undetectable HLA-C antigens, we have established a DNA-based tissue typing method for the HLA-C locus by PCR-SSP (polymerase chain reaction-sequence specific primers). Genomic DNA prepared from Iymphoblastoid 21 B-cell lines and 120 Korean individuals by proteinase K digestion and pheno/chloroform extractions have been typed by PCR-SSP (23 primer mixes were used). The PCR-SSP results of control cell lines were discrepant from serology in 1 case among 21 cases: Cw6 which was negative by serology but positive by PCR-SSP (cell line: MANIKA). Twenty four HLA-Cw "blank" antigens among fifty Korean individuals were completely determined by PCR-SSP DNA typing. HLA-Cw*0101 (15.3%), Cw*1401 (12.3%) and Cw*0701 (11.7%) alleles were frequently found in 120 Korean individual samples. In conclusion. the high level of discrimination for HLA-C alleles may prove useful and informative in the study of transplant survival, and identify the importance of allelic differences, not readily detectable by serology, on host and donor compatibility.

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Genotyping of HLA-A by Polymerase Chain Reaction-Sequence Specific Primer (Polymerase Chain Reaction-Sequence Specific Primer를 이용한 HLA-A 유전자의 DNA 다형성 조사)

  • Jang, Soon-Mo
    • Korean Journal of Clinical Laboratory Science
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    • v.40 no.2
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    • pp.94-97
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    • 2008
  • The human leukocyte antigen (HLA) is the name of the major histocompatibility complex (MCH) in humans. The superlocus contains a large number of genes related to immune system function in humans. This group of genes resides on chromosome 6. and encode cell surface antigen-presenting proteins and many other genes. HLA class I antigen (A, B & C) present peptides from inside the cell. These peptides are produced from digested proteins that are broken down in the lysozymes. Most expressed HLA loci exhibit a remarkable degree of allelic polymorphism, which derives from sequence differences predominantly localized to discrete hypervariable regions of the amino terminal domain of the molecule. In this sutdy, the HLA-A genotypes were determined in twenty students unrelated koreans using the PCR-SSP (Polymerase Chain Reaction-Sequence Specific Primer) technique. Several specific primer pairs in assigning the HLA-A gene were used (A*0201, A*33, A*2401). The results of PCR-SSP, the HLA-A*0201 primer was detected eleven (55%), the HLA-A*33 were detected seven (35%) and the HLA-A*2401 were detected seven (35%). This study shows that the PCR-SSP technique is relatively simple, fast and a practical tool for the determination of the HLA-A genotypes.

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DNA Polymorphism Analysis of the HLA-DRB1 Gene Using Polymerase Chain Reaction-Sequence Specific Primer (PCR-SSP) among Korean Subjects

  • Lee, Kyung-Ok;Park, Taek-Kyu;Park, Young-Suk;Oh, Moon-Ju;Kim, Yoon-Jung
    • BMB Reports
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    • v.29 no.1
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    • pp.45-51
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    • 1996
  • Most expressed HLA loci exhibit a remarkable degree of allelic polymorphism, which derives from sequence differences predominantly localized to discrete hypervariable regions of the amino-terminal domain of the molecule. In this study, the HLA-DRB1 genotypes were determined in eighteen control cell lines and 112 unrelated Koreans using the PCR-SSP (Polymerase Chain Reaction-Sequence Specific Primer) technique. 29 specific primer pairs in assigning the DRB1 gene were used. The results of control cells correlated well with the data which was previously reported. The heterozygosity and homozygosity of the DRB1 gene were 0.786 and 0.214, respectively. In a total of 41 different DRB1 alleles and 83 genotypes, the most frequent allele and genotype were DRB1*04 and DRB1*0901/1501, respectively. This study shows that the PCR-SSP technique is relatively simple, fast and a practical tool for the determination of the HLA-DRBI genotypes. Moreover, these results-allele and genotype frequency and heterozygosity of the HLA DRB1 gene-could be useful for database study before being applied to individual identification and transplantation immunity.

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A PCR Method to Distinguish Matsumuraeses phaseoli from M. falcana Based on the Difference of Nucleotide Sequence in the Mitochondrial Cytochrome c Oxidase Subunit I (미토콘드리아 COI 영역의 뉴클레오티드 서열 차이를 이용한 팥나방과 어리팥나방의 PCR 판별법)

  • Seo, Bo Yoon;Jung, Jin Kyo;Cho, Jum Rae;Kim, Yonggyun;Park, Chang Gyu
    • Korean journal of applied entomology
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    • v.51 no.4
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    • pp.365-370
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    • 2012
  • The two closely related major leguminous crop pests in Korea, Matsumuraeses phaseoli and M. falcana (Lepidoptera: Tortricidae) have very similar morphological characters, which occasionally give rise to a failure in distinguishing between the two. In this study, we report an easy PCR-SSP method to distinguish between them, with a sequence specific primer set (P-SF2, F-SF3, and C-SR3) based on single nucleotide mismatch in 3' terminal base of a primer, which is found in the mitochondrial cytochrome c oxidase subunit I DNA (mtCOI). Through application of this method, each species may be clearly identified in terms of its PCR band size and pattern, only one band (245 bp) for M. falcana and one (409 bp) or two bands (409 bp & 245 bp) for M. phaseoli.

Genetic Distance of Allium Section Cepa by DNA Fingerprint

  • Kim, Haeng-Hoon;Cho, Eun-Gi;Baek, Hyung-Jin;Kim, Chang-Yung;Chae, Young-Am
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.48 no.1
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    • pp.31-37
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    • 2003
  • Identification of compatible parental line is of great importance in introduction of useful characters to onion breeding program, beyond the severe hybridization barrier. Phylogenic analysis of Allium section Cepa was conducted through PCR by URPs, repeated sequences of A. fistulosum, and microsatellite markers. Totally 76 accessions originated from 21 countries were clustered into five groups at a 0.84-similarity level: group I;A. cepa and its wild relatives and A. cepa ssp. ascalonicum, group II; A. cepa ssp. wakegii, A. cepa ssp. proliferum and Samcheung-pa group III; A. fistulosum and A. altaicum, group IV; A. galanthum, group V; Soeckkori-pa. Samcheung-pa and Soekkori-pa, Korean local varieties, shared band type of both Cepa group and Altaicum group, indicating that those are derived from interspecific hybridization between A. fistulosum and A. cepa.

Development of an Effective PCR Technique for Analyzing T-DNA Integration Sites in Brassica Species and Its Application (배추과에서 T-DNA 도입 위치 분석을 위한 효과적인 PCR 방법 개발 및 이용)

  • Lee, Gi-Ho;Yu, Jae-Gyeong;Park, Young-Doo
    • Horticultural Science & Technology
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    • v.33 no.2
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    • pp.242-250
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    • 2015
  • Insertional mutagenesis induced by T-DNA or transposon tagging offers possibilities for analysis of gene function. However, its potential remains limited unless good methods for detecting the target locus are developed. We describe a PCR technique for efficient identification of DNA sequences adjacent to the inserted T-DNA in a higher plant, Chinese cabbage (Brassica rapa ssp. pekinensis). This strategy, which we named variable argument thermal asymmetric interlaced PCR (VA-TAIL PCR), was designed by modifying a single-step annealing-extension PCR by including a touch-up PCR protocol and using long gene-specific primers. Amplification efficiency of this PCR program was significantly increased by employing an autosegment extension method and linked sequence strategy in nested long gene-specific primers. For this technique, arbitrary degenerate (AD) primers specific to B. rapa were designed by analyzing the Integr8 proteome database. These primers showed higher accuracy and utility in the identification of flanking DNA sequences from individual transgenic Chinese cabbages in a large T-DNA inserted population. The VA-TAIL PCR method described in this study allows the identification of DNA regions flanking known DNA fragments. This method has potential biotechnological applications, being highly suitable for identification of target genomic loci in insertional mutagenesis screens.