• Title/Summary/Keyword: AFLP clones

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Application of Chromosome Manipulation, DOP-PCR and AFLP Methods to Isolate Sex-Specific DNAs from Rumex acetosa L.

  • Jin, Dong-Chung;Kim, Joong-Soon;Park, ji-Young;Bong, Jae-Wook;Hur, Yoon-Kang
    • Journal of Photoscience
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    • v.12 no.2
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    • pp.75-82
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    • 2005
  • Rumex acetosa L. is a dioecious flowering plant with well developed sex chromosome system: 2n = 12 + XX in the female plants and 2n = 12 + XY1Y2 in the male plants. To isolate sex-linked DNA, we carried out chromosome micromanipulation, followed by DOP-PCR, AFLP of the PCR products, reverse Southern hybridization and sequence analysis. From 500 AFLP specific clones, 13 X-chromosome and 5 Y-chromosome specific clones were obtained. Except one clone RADAX-239 ($\underline{R}umex\;\underline{a}-\underline{D}OP-PCR-\underline{A}FLP-\underline{Y}-chromosome\;specific$), all clones appear to be R. acetosa plant-specific sequences and non-coding sequences. Southern blot analysis using these clones could not discriminate genomic DNAs either from male or female plants. Results of this study imply that both autosome-origin and degeneration of sex chromosomes are prevalent in plant systems.

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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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Combined Genome Mapping of RFLP-AFLP-SSR in Pepper

  • Lee, Je Min;Kim, Byung-Dong
    • Genomics & Informatics
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    • v.1 no.2
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    • pp.108-112
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    • 2003
  • We have constructed a molecular linkage map of pepper (Capsicum spp.) in an interspecific $F_2$ population of 107 plants with 320 RFLP, 136 AFLP, and 46 SSR markers. The resulting linkage map consists of 15 linkage groups covering 1,720 cM with an average map distance of 3.7 cM between framework markers. Most RFLP markers ($80\%$) were pepper-derived clones and these markers were evenly distributed all over the genome. Genes for defense and biosynthesis of carotenoids and capsaicinoids were mapped on this linkage map. By using 30 primer combinations, AFLP markers were generated in the $F_2$ population. For development of SSR markers in Capsicum, microsatellites were isolated from two small-insert genomic libraries and the GenBank database. This combined map provides a starting point for high-resolution QTL analysis, gene isolation, and molecular breeding.

Linkage Map Construction and Molecular Genetic Approach in Capsicum spp.

  • Kim, Byung-Dong
    • Korean Journal of Plant Tissue Culture
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    • v.27 no.5
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    • pp.367-373
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    • 2000
  • We have constructed a molecular linkage map of chili pepper (Capsicum spp.) in an interspecific (C. annuum cv. TF68 x C. chinense cv. Habanero) F$_2$ population of 107 plants with 150 RFLP and 430 AFLP markers. The resulting linkage map consists of 11 large (206-60.3 cM) and 5 small (32.6- 10.3 cM) linkage groups cover-ing 1,320 cM with an average map distance between framework markers of 7.5 cM. Most (80%) of the RFLP markers were pepper-derived clones and these markers were evenly distributed across the genome. By using 30 primer combinations, 444 AFLP markers were generated in the F$_2$population. The majority of the AFLP markers clustered in each linkage group, although PstI/MseI markers were more evenly distributed than Eco RI/MseI markers within the linkage groups. Genes for biosynthesis of carotenoids and capsaicinoids were mapped on our linkage map. This map will provide the basis of studying secondary metabolites in pepper.

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Genetic Diversity among Dioscorea, spp. Using Molecular Markers (분자표지를 이용한 마(Dioscorea spp.)의 다양성 분석)

  • Chang, K.J.;Choi, I.Y.;Park, J.H.;Park, J.I.;Yoon, B.S.;Kim, N.S.
    • Journal of Practical Agriculture & Fisheries Research
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    • v.4 no.1
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    • pp.45-54
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    • 2002
  • A lot of clones of the genus Dioscorea species have been introduced from some tropical and subtropical regions since 1997. The aim of this study was to determine characteristics of the production process in yams(Dioscorea spp). By utilizing 51clones of water yams(Dioscorea spp), some morphological characteristics were investigated at the field. Intraspecific genetic relationship of 51 variation types of the Yam classified by their external morphological characteristics such as leaf and tuber shape were assessed by DNA using random and specific primers. D. Alata and D. opposita were showed different relationship between yield and the growth of their aboveground parts. This suggests that even in the same species there were differences in yield volume and translocation of assimilation products, depending on the types. D. alata were distinguished from others Dioscorea species at 62% level in AFLP analysis. Also in principal component analysis, D. alata were showed the class from II to V.

Identification of Differentially Expressed Genes by cDNA-AFLP in Magnaporthe oryzae

  • Chi, Myoung-Hwan;Park, Sook-Young
    • Research in Plant Disease
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    • v.25 no.4
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    • pp.205-212
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    • 2019
  • Analysis of differentially expressed genes has assisted discovery of gene sets involved in particular biological processes. The purpose of this study was to identify genes involved in appressorium formation in the rice blast fungus Magnaporthe oryzae via analysis of cDNA-amplified fragment length polymorphisms. Amplification of appressorial and vegetative mycelial cDNAs using 28 primer combinations generated over 200 differentially expressed transcript-derived fragments (TDFs). TDFs were excised from gels, re-amplified by PCR, cloned, and sequenced. Forty-four of 52 clones analyzed corresponded to 42 genes. Quantitative real-time PCR showed that expression of 23 genes was up-regulated during appressorium formation, one of which was the MCK1 gene that had been shown to be involved in appressorium formation. This study will be providing valuable resources for identifying the genes such as pathogenicity-related genes in M. oryzae.

Progress and Prospect of Rice Biotechnology in Korea

  • Tae Young, Chung
    • Proceedings of the Korean Society of Sericultural Science Conference
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    • 1997.06a
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    • pp.23-49
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    • 1997
  • This is a progress report of rice biotechnology including development of gene transformation system, gene cloning and molecular mapping in rice. The scope of the research was focused on the connection between conventional breeding and biotech-researches. Plant transformation via Agrobacterium or particle bombardment was developed to introduce one or several genes to recommended rice cultivars. Two chimeric genes containing a maize ribosome inactivating protein gene (RIP) and a gerbicide resistant gene (bar) were introduced to Nipponbare, a Japonica cultivar, and transmitted to Korean cultivars. The homozygous progenies of herbicide resistant transgenic plant showed good fertility and agronomic characters. To explore the genetic resourses in rice, over 8,000 cDNA clones from immature rice seed have been isolated and sequenced. About 13% of clones were identified as enzymes related to metabolic pathway. Among them, twenty clones have high homology with genes encoding enzymes in the photorespiratory carbon cycle reaction. Up to now about 100 clones were fully sequenced and registered at EMBL and GenBank. For the mapping of quantitative tarits loci (QTL) and eternal recombinant inbred population with 164 F13 lines (MGRI) was developed from a cross between Milyang 23 and Gihobyeo, Korean rice cultivars. After construction of fully saturated RFLP and AFLP map, quantitative traits using MGRI population were analyzed and integrated into the molecular map. Eighty seven loci were determined with 27 QTL characters including yield and yield components on rice chromosomes. Map based cloning was also tried to isolate semi-dwarf (sd-1) gene in rice. A DNA probe, RG 109, the most tightly linked to sd-1 gene was used to screen from bacterial artifical chromosome (BAC) libraries and five over lapping clones presumably containing sd-1 gene were isolated. Rice genetic database including results of biotech reasearch and classical genetics is provided at Korea Rice Genome Server which is accessible with world wide web (www) browser. The server provides rice cDNA sequences and map informations linked with phenotypic images.

Construction of an Integrated Pepper Map Using RFLP, SSR, CAPS, AFLP, WRKY, rRAMP, and BAC End Sequences

  • Lee, Heung-Ryul;Bae, Ik-Hyun;Park, Soung-Woo;Kim, Hyoun-Joung;Min, Woong-Ki;Han, Jung-Heon;Kim, Ki-Taek;Kim, Byung-Dong
    • Molecules and Cells
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    • v.27 no.1
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    • pp.21-37
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    • 2009
  • Map-based cloning to find genes of interest, marker-assisted selection (MAS), and marker-assisted breeding (MAB) all require good genetic maps with high reproducible markers. For map construction as well as chromosome assignment, development of single copy PCR-based markers and map integration process are necessary. In this study, the 132 markers (57 STS from BAC-end sequences, 13 STS from RFLP, and 62 SSR) were newly developed as single copy type PCR-based markers. They were used together with 1830 markers previously developed in our lab to construct an integrated map with the Joinmap 3.0 program. This integrated map contained 169 SSR, 354 RFLP, 23 STS from BAC-end sequences, 6 STS from RFLP, 152 AFLP, 51 WRKY, and 99 rRAMP markers on 12 chromosomes. The integrated map contained four genetic maps of two interspecific (Capsicum annuum 'TF68' and C. chinense 'Habanero') and two intraspecific (C. annuum 'CM334' and C. annuum 'Chilsungcho') populations of peppers. This constructed integrated map consisted of 805 markers (map distance of 1858 cM) in interspecific populations and 745 markers (map distance of 1892 cM) in intraspecific populations. The used pepper STS were first developed from end sequences of BAC clones from Capsicum annuum 'CM334'. This integrated map will provide useful information for construction of future pepper genetic maps and for assignment of linkage groups to pepper chromosomes.