• Title/Summary/Keyword: direct sequencing

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Genetic Stock Identification of Common Carp (Cyprinus carpio) by Detection of Intraspecific DNA Sequence Variation in the Mitochondrial 12S rRNA Gene (미토콘드리아 12S rRNA 유전자 변이 조사를 통한 잉어(Cyprinus carpio)의 유전학적 동정)

  • 남윤권;주수동;정창화;노충환;조재윤;김동수
    • Journal of Aquaculture
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    • v.10 no.4
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    • pp.403-407
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    • 1997
  • Intraspecific sequence variation was detected by polymerase chain reaction (PCR) and direct sequencing of a 350-nucleotide region of the mitochondrial 12S rRNA gene of two natural populations (Han River and Nakdong River) and one hatchery stock (Jinhae Inland Fisheries Institute) of local strain common carp, one Israeli strain of common carp stock from Pukyong National University (PKU), and one hybrid between Israeli strain of common carp female and local strain common carp male from PKU stock. There is little variation in 350 bases of the mitochondrial 12S rRNA gene sequences among 2 natural and 1 hatchery local strain common carp populatins, representing abut 7 to 20 nucleotide differences (less than 6%). The sequence of specimens from Han River was more similar to that from Nakdong River (identity=98.0%) than to that from Jinhae Inland Fisheries Institute (identity=96.3%). Sequence variation between Israeli strain and wild local strain common carp was higher than the variation within natural stocks. The level of variation was ranged from 15.7 to 17.7%. The hybrid showed very similar nucleotide4 sequence of 12S rRNA gene to the sequence of Israeli strain with the identity of 98.9%.

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Phylogeny of the subfamily Salmoninae distributed in Korea based upon nucleotide sequences of mitochondrial ribosomal RNA genes (미토콘드리아 ribosomal RNA 유전자 염기서열분석에 의한 한국산 연어아과 어류의 유전적 계통도)

  • LEE Heui-Jung;PARK Jung-Youn;LEE Jeong-Ho;MIN Kwang-Sik;JEON Im Gi;YOO Mi-Ae;LEE Won-Ho
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.33 no.2
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    • pp.103-109
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    • 2000
  • Complete senuences of the mitochondrial rRNA Benes were determined among six salmonines in Korean Waters (Brachpmystax lenok, Onoorhpchus keta, O. masou mason, O. mason ishikawae, O. mykiss, and albino mutant of O. mykiss). The purposes of this study were to provide the basic information on levels of mtDNA polymorphism among these species for genetic characterization; discuss phylogentic relationships among three Oncorhynchus sepecies; demonstrate the utility of rRNA gene sequence data as a genetic marker for disringuishinf among Korean salmonines. PCR/direct sequencing data indicated the following consistent results; 1) 12S rRNA genes was 945 bases long in Oncorhynchus species, and 946 bases in B. lenot including one insertion. 2) Of sequence variation in mitochondrial rRNA regions, transitional substitutions were superior to transversion. 3) The significant differences were not shown in the intraspecific variation values in these gene regions. The percentage sequence divergence values were ranged from $0.066 to 0.212{\%}$. 4) The interspecific divergences were greater than the intraspecific variation. Nevertheless, ribosomal RMh genes were more conserved among species than the other mitochondrial genes, and they showed potentiality as an intergenic marker for systematics. In addition, phylogenetic trees, constructed from this data, supported that cherry salmon was closer to chum salmon than to rainbow trout, and that lenok was most distantly related species in six salmonid species.

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Detection of Mutations to Zidovudine in the pol Gene of Human Immunodeficiency Virus-1 by Direct Sequencing (인면역결핍 바이러스 pol 유전자 염기서열 결정에 의한 지도부딘 (ZDV) 내성 돌연변이의 탐지)

  • Cho, Young-Keol;Lee, Hee-Jung;Sung, Heung-Sup;Kim, Yoo-Kyum;Kim, Young-Bong;Lee, Yong-Jin;Kim, Mi-Jung;Kim, Dae-Ghon;Won, Young-Ho;Cho, Goon-Jae
    • The Journal of Korean Society of Virology
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    • v.29 no.4
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    • pp.271-281
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    • 1999
  • The nested polymerase chain reaction (PCR) assay was used to determine the sequences of reverse transcriptase (RT) codons 41, 67, 70, 210, 215 and 219 of human immunodeficiency virus-1 (HIV-1) pol gene. Template DNA was obtained from uncultured peripheral blood mononuclear cells from 27 Korean HIV-1 infected patients treated with ZDV and Korean red ginseng. The second PCRs were done for 2 separated regions (RT codons $13{\sim}98$ and $152{\sim}259$) with $5\;{\mu}l$ of the first PCR productNucleotide sequences were determined by direct sequencing. In the 27 patients, CD4+ cell count decreased from $230{\pm}117/{\mu}l$ to $152{\pm}162/{\mu}l$ for $46{\pm}26$ months (Mo), and actual duration of ZDV intake was $72{\pm}16$ Mo. In the 16 patients who had been treated with ZDV therapy ${\ge}25$ Mo, the incidences of 70R, 215F/Y, and 41L were 61%, 28% and 22%, respectively and those of 67N, 210W and 219Q were 17%. The incidences of 215F/Y were 6.7% for group ${\le}12$ Mo treatment, 22.7% for group with 13 to 24 Mo, and 27.8% for group ${\ge}25$ Mo. There was no mutation in 9 patients. It might be associated with the interruption of ZDV therapy for more than 6 months in 6 patients. This study shows that the detection of mutation could be useful prognostic marker with other clinical and virological data, and very low mutation rate is dectected compared to overseas reports.

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A genetic algorithm for determining the optimal operating policies in an integrated-automated manufacturing system (통합자동생산시스템에서 최적운영방안 결정을 위한 유전자 알고리즘의 개발)

  • 임준묵
    • Proceedings of the Korea Society for Industrial Systems Conference
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    • 1999.05a
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    • pp.145-153
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    • 1999
  • We consider a Direct Input Output Manufacturing System(DIOMS) which has a munber of machine centers placed along a built-in Automated Storage/Retrieval System(AS/RS). The Storage/Retrieval (S/R) machine handles parts placed on pallets for the machine centers located at either one or both sides of the As/Rs. This report studies the operational aspect of DIOMS and determines the optimal operating policy by combining computer simulation and genetic algorithm. The operational problem includes: input sequencing control, dispatching rule of the S/R machine, machine center-based part type selection rule, and storage assignment policy. For each operating policy, several different policies are considered based on the known research results. In this report, using the computer simulation and genetic algorithm we suggest a method which gives the optimal configuration of operating policies within reasonable computation time.

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Detection of Rifampin Resistance Mutation and Its Altered Nucleotide Sequences in Mycobacterium leprae Isolated from Korean Patients with Leprosy

  • Kim, Soon-Ok;Kim, Min-Joo;Tae, Chae-Gue;Suh, Joo-Won
    • Journal of Microbiology
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    • v.34 no.3
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    • pp.236-240
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    • 1996
  • Rifampin is the most powerful drug for treating leprosy and tuberculosis today. It inhibits initiation and elongation of RNA transcription by binding to $\beta$-subunit of RNA polymerase, leading to kill mycobacteria. We isolated one variant strain of Mycobacterium leprae from 24 Korean leprosy patients who are less susceptible to rifampin or have suffered from relapse by polymerase chain reaction and single strand conformation polymorphism (PCR-SSCP) of the rpoB gene. Direct sequencing of the rpoB region of M. leprae variant revealed missense mutations which altered the amino acids sequenceof RpoB to Ser-464, Arg-465, Arg-467 and Ala-468. This is the first finding on rpoB gene mutation of M. leprae from Korean patients ; moreover the mutant type was found to be different from the previously reported cases in other countries.

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Lack of Mutation in p53 and H-ras Genes in Phenytoin Induced Gingival Overgrowth Suggests its Non Cancerous Nature

  • Jayaraman, Bhaskar;Valiathan, Gopalakrishnan Mohan;Jayakumar, Keerthivasan;Palaniyandi, Arunmozhi;Thenumgal, Siji Jacob;Ramanathan, Arvind
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.11
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    • pp.5535-5538
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    • 2012
  • Background: There have been case reports of oral squamous cell carcinoma arising from gingival overgrowth induced by phenytoin - an antiepileptic drug. However, a detailed analysis for the presence of mutations in p53 and ras genes, which are the two most frequently mutated genes in cancers, in phenytoin induced gingival overgrowth tissues has hitherto not been performed. Methods: Cellular DNA isolated from twenty gingival overgrowth tissues collected from patients undergoing phenytoin therapy were amplified using primers for p53 (exons 5-8) and H-ras (exons 1-2) genes. The PCR amplicons were then gel purified and subjected to direct sequencing analysis to screen for mutations. Results: Direct sequencing of twenty samples of phenytoin induced gingival growth did not identify mutations in any of the exons of p53 and H-ras genes that were analyzed. Conclusion: Our result indicates that mutational alteration of p53 and H-ras genes is infrequent in phenytoin induced gingival growth, which thus suggests a non malignant nature of this pathology. The findings in the present study are clinically significant as a large number of epileptic patients are treated with phenytoin.

The origin-of-cell harboring cancer-driving mutations in human glioblastoma

  • Lee, Joo Ho;Lee, Jeong Ho
    • BMB Reports
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    • v.51 no.10
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    • pp.481-483
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    • 2018
  • Glioblastoma (GBM) is the most common and aggressive form of human adult brain malignancy. The identification of the cell of origin harboring cancer-driver mutations is the fundamental issue for understanding the nature of GBM and developing the effective therapeutic target. It has been a long-term hypothesis that neural stem cells in the subventricular zone (SVZ) might be the origin-of-cells in human glioblastoma since they are known to have life-long proliferative activity and acquire somatic mutations. However, the cell of origin for GBM remains controversial due to lack of direct evidence thereof in human GBM. Our recent study using various sequencing techniques in triple matched samples such as tumor-free SVZ, tumor, and normal tissues from human patients identified the clonal relationship of driver mutations between GBM and tumor-free SVZ harboring neural stem cells (NSCs). Tumor-free SVZ tissue away from the tumor contained low-level GBM driver mutations (as low as 1% allelic frequency) that were found in the dominant clones in its matching tumors. Moreover, via single-cell sequencing and microdissection, it was discovered that astrocyte-like NSCs accumulating driver mutations evolved into GBM with clonal expansion. Furthermore, mutagenesis of cancer-driving genes of NSCs in mice leads to migration of mutant cells from SVZ to distant brain and development of high-grade glioma through the aberrant growth of oligodendrocyte precursor lineage. Altogether, the present study provides the first direct evidence that NSCs in human SVZ is the cell of origin that develops the driver mutations of GBM.

Screening of BRCA1/2 Mutations Using Direct Sequencing in Indonesian Familial Breast Cancer Cases

  • Anwar, Sumadi Lukman;Haryono, Samuel J;Aryandono, Teguh;Datasena, I Gusti Bagus
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.4
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    • pp.1987-1991
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    • 2016
  • Breast cancer has emerged as the most prevalent cancer among women worldwide, including in Indonesia. The contribution of genes associated with high-risk breast-ovarian cancers, BRCA1 and BRCA2, in the Indonesian population is relatively unknown. We have characterized family history of patients with moderate- to high-risk of breast cancer predisposition in 26 unrelated cases from Indonesia for BRCA1/2 mutation analyses using direct sequencing. Known deleterious mutations were not found in either BRCA1 or BRCA2 genes. Seven variants in BRCA2 were documented in 10 of 26 patients (38%). All variants were categorized as unclassified (VUSs). Two synonymous variants, c.3623A>G and c.4035T>C, were found in 5 patients. One variant, c4600T>C, was found in a 38 year old woman with a family history of breast cancer. We have found 4 novel variants in BRCA2 gene including c.6718C>G, c.3281A>G, c.10176C>G, and c4490T>C in 4 unrelated patients, all of them having a positive family history of breast cancer. In accordance to other studies in Asian population, our study showed more frequent variants in BRCA2 compared to BRCA1. Further studies involving larger numbers of hereditary breast cancer patients are required to reveal contribution of BRCA1/2 mutations and/or other predisposing genes among familial breast cancer patients in Indonesia.

Identification of quantitative trait loci for root development during seedling stage in rice

  • Han, Jae-Hyuk;Chin, Joong Hyoun;Yoo, Soo-Cheul
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2017.06a
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    • pp.103-103
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    • 2017
  • Vigorous root growth at the seedling stage in dry direct-seeded conditions is considered as a critical trait because it is involved in seedling emergence, early vegetative vigour, nutrient uptake as well as drought tolerance. In this study, we performed QTL mapping using the recombinant inbred lines obtained from the cross between Tongil-type Dasan and temperate japonica TR22183 (DT-RILs) to identify QTL underlying early root development. TR22183, which was previously reported to have high nitrogen utility and cold tolerance, showed vigorous root growth at the seedling stage in semi-drought conditions. Root length, fresh weight and dry weight of TR22183 were significantly higher than in Dasan. By QTL analysis with genotyping-by-sequencing method, we identified two QTLs for root fresh weight (RFW) in chromosome 7 and root dry weight (RDW) in chromosome 8, explaining phenotypic variances of 13.5% and 10.6%, respectively. These QTLs would be used to develop rice varieties adapted to direct-seeded cultivating system.

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Reflections on the US FDA's Warning on Direct-to-Consumer Genetic Testing

  • Yim, Seon-Hee;Chung, Yeun-Jun
    • Genomics & Informatics
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    • v.12 no.4
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    • pp.151-155
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    • 2014
  • In November 2013, the US Food and Drug Administration (FDA) sent a warning letter to 23andMe, Inc. and ordered the company to discontinue marketing of the 23andMe Personal Genome Service (PGS) until it receives FDA marketing authorization for the device. The FDA considers the PGS as an unclassified medical device, which requires premarket approval or de novo classification. Opponents of the FDA's action expressed their concerns, saying that the FDA is overcautious and paternalistic, which violates consumers' rights and might stifle the consumer genomics field itself, and insisted that the agency should not restrict direct-to-consumer (DTC) genomic testing without empirical evidence of harm. Proponents support the agency's action as protection of consumers from potentially invalid and almost useless information. This action was also significant, since it reflected the FDA's attitude towards medical application of next-generation sequencing techniques. In this review, we followed up on the FDA-23andMe incident and evaluated the problems and prospects for DTC genetic testing.