• 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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    • v.7 no.2
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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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    • v.34 no.2
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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.

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

  • Yoon, Young-Eun;Park, Sang-Yong;Bae, Joo-Seung;Bang, In-Chul
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.42 no.2
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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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    • 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.

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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    • v.22 no.9
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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.

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

  • Park, Cho Rong;Kim, Jeong Hee;Yu, Yong Man;Youn, Young Nam
    • Korean journal of applied entomology
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    • v.55 no.3
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    • pp.245-256
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    • 2016
  • Elytra of Harmonia axyridis exhibit varied color patterns. In the present study, we deciphered the genetic basis for intraspecific diversity of elytra color patterns in H. axyridis, using amplified fragment length polymorphism (AFLP). Twenty-eight AFLP reactions were performed to generate a total of 2,741 bands. Of these, 20 bands were polymorphic for each color pattern. The polymorphic bands showed differences of genetic character among different color patterns of H. axyridis. Among them, ten candidate AFLP markers were color-linked. S1, S2, and S20 markers were detected in Succinea 1 and 2 variants of H. axyridis, whereas S3 and S5 were specifically detected in the Conspicua variant. S15, S18, and S19 were specific to the Succinea 2 variant. Polymerase chain reaction (PCR) products of these ten AFLP markers were sequenced. BLAST analysis of these sequences against the GenBank database revealed their homology to DNA fragments of unknown function. Based on the color-linked AFLP markers, sequence characterized amplified region (SCAR) markers were designed for PCR amplification of genomic DNA. Of the ten AFLP markers, five were successfully converted into SCAR markers, which could discriminate elytra color polymorphism in H. axyridis.

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.

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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    • v.17 no.4
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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.