• Title/Summary/Keyword: amplified fragment length polymorphism

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Cloning and Characterization of Genes Controlling Flower Color in Pharbitis nil Using AFLP (Amplified Fragment Length Polymorphism) and DDRT (Differential Display Reverse Transcription)

  • Kim, Eun-Mi;Jueson Maeng;Lim, Yong-Pyo;Yoonkang Hur
    • Journal of Photoscience
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    • 제7권2호
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    • pp.73-78
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    • 2000
  • To analyze molecular traits determining pigmentation between Pharbitis nill violet and white, Amplified Fragment Length Polymorphism(AFLP) and Differential Display Reverse Transcription(DDRT) experiments were carried out with either genomic DNAs or total RNAs isolated from both plants. Results of AFLP experiment in combination of 8 EcoRⅠ primers with 6 MseⅠ primers showed 41 violet-and 60 white-specific DNA bands. In the subsequent experiment, 22 violet-and 22 white-specific DNA fragments were amplified by PCR with DNAs eluted. The sizes of the fragments range from 200 to 600bp. DDRT using total RNA produced 19 violet-and 17 white-specific cDNA fragments, ranging from 200 to 600bp. The fragments obtained by both AFLP and DDRT had been cloned into pGEM T-easy vector, amplified and subjected to the nucleotide sequence analyses. As a result of Blast sequence analysis, most of them sequenced up to date showed no similarity to any Known gene, while few has similarity to known animal or plant genes. An AFLP clone V6, for example, has a strong sequence similarity to the human transcription factor LZIP-alpha mRNA and a DDRT clone W19 to Solanum tuberosum 3-hydroxy-3-methylglutaryl coenzyme A reductase mRNA.

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Comparison of RAPD, AFLP, and EF -1 α Sequences for the Phylogenetic Analysis of Fusarium oxysporum and Its formae speciales in Korea

  • Park, Jae-Min;Kim, Gi-Young;Lee, Song-Jin;Kim, Mun-Ok;Huh, Man-Kyu;Lee, Tae-Ho;Lee, Jae-Dong
    • Mycobiology
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    • 제34권2호
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    • pp.45-55
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    • 2006
  • Although Fursarium oxysporum causes diseases in economically important plant hosts, identification of F. oxysporum formae speciales has been difficult due to confusing phenotypic classification systems. To resolve these complexity, we evaluated genetic relationship of nine formae speciales of F. oxysporum with random amplified polymorphic DNA (RAPD), amplified fragment length polymorphism (AFLP), and translation elongation factor-l alpha ($EF-1{\alpha}$) gene. In addition, the correlation between mycotoxin content of fusaric acid and isolates based on molecular marker data was evaluated using the modified Mantel's test. According to these result, these fusaric acid-producing strains could not identify clearly, and independent of geographic locations and host specificities. However, in the identification of F. oxysporum formae speciales, especially, AFLP analysis showed a higher discriminatory power than that of a the RAPD and $EF-1{\alpha}$ analyses, all three techniques were able to detect genetic variability among F. oxysporum formae speciales in this study.

AFLP 분석에 의한 병어속 (Pampus) 3종의 유전 변이 (Genetic Variation of the Three Pampus spp. (Pisces: Stromateidae) using Amplified Fragment Length Polymorphism (AFLP))

  • 윤영은;박상용;배주승;방인철
    • 한국수산과학회지
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    • 제42권2호
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    • pp.146-150
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    • 2009
  • Genetic variation and relationship of two wild (Pampus argenteus and P. echinogaster) and one cultured (P. chinesis) pomfret fish belonging to the genus Pampus were assessed. Specimens were collected from Korea and China and subjected to amplified fragment length polymorphism (AFLP) DNA fingerprinting. Four primer combinations generated a total of 304 DNA fragments ranging from 153 to 251 bands. Polymorphism and genetic diversity of cultured P. chinensis (22.9% and 0.038) were significantly lower than the two wild species of P. argenteus (93.6% and 0.311) and P. echinogaster (94.0% and 0.290). Genetic distance ranged from 0.335 (P. argenteus and P. echinogaster) to 0.646 (P. argenteus and P. chinensis) and showed a congeneric relationship within this genus. Twenty one of specific AFLP markers from four primer combinations bands were produced. These results suggest that AFLP polymorphism may be a useful marker for genetic identification among the three species studies here.

Authentication of Salted-dried Fish Species Using Polymerase Chain Reaction-Single Strand Conformational Polymorphism and Restriction Analysis of Mitochondrial DNA

  • Kim, Joo-Shin;Chu, Kin Kan Astley;Kwan, Hoi Shan;Chung, Hau Yin
    • 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.

A Simple Polymerase Chain Reaction-based Method for the Discrimination of Three Chicken Breeds

  • Kubo, Y.;Plastow, G.;Mitsuhashi, Tadayoshi
    • Asian-Australasian Journal of Animal Sciences
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    • 제22권9호
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    • pp.1241-1247
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    • 2009
  • A large number of branded chicken products exist in Japan, and in some cases, the breed of chicken is an important factor used to attract consumer interest in the retail product. In order to establish a simple method for verifying such breed claims we applied the amplified fragment length polymorphism (AFLP) technique to nine chicken breeds (White Cornish, Red Cornish, White Plymouth Rock, New Hampshire, Rhode Island Red, Barred Plymouth Rock, Hinaidori, Tosajidori, Tsushimajidori) to search for molecular markers able to discriminate chicken breeds. Three breed-specific single nucleotide polymorphisms (SNP) were identified, one for each of Hinaidori, Tosajidori, or New Hampshire. A total of 219 individuals from the nine breeds were analyzed using a specific PCR test for each of these SNP. The PCR tests made it possible to discriminate between the breeds of chickens to identify products from these three breeds. This PCR method provides an efficient method for the routine analysis and verification of certified chicken products.

Amplified Fragment Length Polymorphism (AFLP)을 이용한 무당벌레(Harmonia axyridis : Coccinellidae)의 초시색상패턴의 변이 분석 (Differentiation of Elytra Color Patterns in Multicolored Asian Ladybird Beetle, Harmonia axyridis (Coleoptera; Coccinellidae), using AFLP analyses)

  • 박초롱;김정희;유용만;윤영남
    • 한국응용곤충학회지
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    • 제55권3호
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    • pp.245-256
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    • 2016
  • 무당벌레(Harmonia axyridis)는 종내에서 초시색상패턴이 매우 다양하게 존재한다. 본 논문에서는 서로 다른 색상패턴의 무당벌레를 대상으로 amplified fragment length polymorphism (AFLP)을 실시하여 무당벌레의 초시색상 패턴간 유전형질의 차이를 확인하고자 하였다. 총 28 개의 프라이머 조합으로 실험을 실시한 결과, 총 2,741 개의 밴드가 검출되었다. 그 중 20 개의 밴드(S1-S20)만이 특정 색상패턴에서 나타났다. 이들 가운데 9 개의 밴드를 색상에 연관된 AFLP 후보 지표로 선발하였다. 밴드 가운데 S1과 S2, S20은 Succinea 1, 2 변이형에 공통적으로 나타났으며, S3와 S5는 Conspicua 변이형에 특이적이었다. 또한 S13는 Spectabilis 변이형에, S15와 S18, S19는 Succinea 2 변이형에 특이적이었다. 특정 색상패턴에만 나타나는 9 개의 AFLP 지표들은 cloning을 통해 염기서열 분석을 실시하였고, GenBank를 이용해 다른 염기 서열과 비교를 해보았지만 아무런 상동성도 찾을 수가 없었다. 무당벌레 종 내 유전적 다양성을 평가한 결과, Spectabilis가 Conspicua보다 Succinea 변이형에 높은 유사성을 보였다. 색상에 연관된 AFLP 후보 지표를 기준으로 sequence characterized amplified region (SCAR) 지표로 변환하여 9 개의 AFLP 분자지표들 가운데에서 5 개만이 SCAR 지표로 전환될 수 있었으며, 이를 통해 AFLP 지표가 무당벌레의 색상과 연관되어 있는지 확인할 수 있었다.

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

  • 김광현;곽길한;송희종;조정곤
    • 한국임상수의학회지
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    • 제27권1호
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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.

Use of Restriction Fragment Length Polymorphism Analysis to Differentiate Fungal Strains in Sunchang Meju

  • Jung, Jong-Hyun;Seo, Dong-Ho;Bhoo, Sung-Hee;Ha, Suk-Jin;Kim, Jong-Sang;Kim, Jeong-Hwan;Kwon, Dae-Young;Cha, Jae-Ho;Park, Cheon-Seok
    • Food Science and Biotechnology
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    • 제17권4호
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    • pp.888-891
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    • 2008
  • Twenty-three fungal strains were isolated from meju that had originated from the Sunchang province, the famous location for making fermented soybean foods in Korea. The restriction fragment length polymorphism (RFLP) of the internal transcribed spacer (ITS) region of the rDNA (ITS-RFLP) was applied to differentiate the isolated fungal strains. First, the ITS region by polymerase chain reaction (PCR) with specific primers was amplified and then cleaved the products with different restriction enzymes. Cleavage of the amplified fragments with the restriction enzymes AluI, HaeIII, HhaI, and TaqI revealed extensive polymorphisms. The ITS-RFLP results highly correlated with ITS sequence analysis. All of the 23 fungal strains were classified into 5 groups by ITS-RFLP analysis. Aspergillus oryzae was the major fungal strain isolated from Sunchang meju (12 out of 23), while Aspergillus fumigatus was the next most frequently isolated strain (7 out of 23). In contrast, it was found that Fusarium asiaticum, Aspergillus sydowii, and Arthrinium sp. were the minor fungal strains in meju.