• Title/Summary/Keyword: NGS

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Development of SNP marker set for marker-assisted backcrossing (MABC) in cultivating tomato varieties

  • Park, GiRim;Jang, Hyun A;Jo, Sung-Hwan;Park, Younghoon;Oh, Sang-Keun;Nam, Moon
    • Korean Journal of Agricultural Science
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    • v.45 no.3
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    • pp.385-400
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    • 2018
  • Marker-assisted backcrossing (MABC) is useful for selecting offspring with a highly recovered genetic background for a recurrent parent at early generation unlike rice and other field crops. Molecular marker sets applicable to practical MABC are scarce in vegetable crops including tomatoes. In this study, we used the National Center for Biotechnology Information- short read archive (NCBI-SRA) database that provided the whole genome sequences of 234 tomato accessions and selected 27,680 tag-single nucleotide polymorphisms (tag-SNPs) that can identify haplotypes in the tomato genome. From this SNP dataset, a total of 143 tag-SNPs that have a high polymorphism information content (PIC) value (> 0.3) and are physically evenly distributed on each chromosome were selected as a MABC marker set. This marker set was tested for its polymorphism in each pairwise cross combination constructed with 124 of the 234 tomato accessions, and a relatively high number of SNP markers polymorphic for the cross combination was observed. The reliability of the MABC SNP set was assessed by converting 18 SNPs into Luna probe-based high-resolution melting (HRM) markers and genotyping nine tomato accessions. The results show that the SNP information and HRM marker genotype matched in 98.6% of the experiment data points, indicating that our sequence analysis pipeline for SNP mining worked successfully. The tag-SNP set for the MABC developed in this study can be useful for not only a practical backcrossing program but also for cultivar identification and F1 seed purity test in tomatoes.

CaGe: A Web-Based Cancer Gene Annotation System for Cancer Genomics

  • Park, Young-Kyu;Kang, Tae-Wook;Baek, Su-Jin;Kim, Kwon-Il;Kim, Seon-Young;Lee, Do-Heon;Kim, Yong-Sung
    • Genomics & Informatics
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    • v.10 no.1
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    • pp.33-39
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    • 2012
  • High-throughput genomic technologies (HGTs), including next-generation DNA sequencing (NGS), microarray, and serial analysis of gene expression (SAGE), have become effective experimental tools for cancer genomics to identify cancer-associated somatic genomic alterations and genes. The main hurdle in cancer genomics is to identify the real causative mutations or genes out of many candidates from an HGT-based cancer genomic analysis. One useful approach is to refer to known cancer genes and associated information. The list of known cancer genes can be used to determine candidates of cancer driver mutations, while cancer gene-related information, including gene expression, protein-protein interaction, and pathways, can be useful for scoring novel candidates. Some cancer gene or mutation databases exist for this purpose, but few specialized tools exist for an automated analysis of a long gene list from an HGT-based cancer genomic analysis. This report presents a new web-accessible bioinformatic tool, called CaGe, a cancer genome annotation system for the assessment of candidates of cancer genes from HGT-based cancer genomics. The tool provides users with information on cancer-related genes, mutations, pathways, and associated annotations through annotation and browsing functions. With this tool, researchers can classify their candidate genes from cancer genome studies into either previously reported or novel categories of cancer genes and gain insight into underlying carcinogenic mechanisms through a pathway analysis. We show the usefulness of CaGe by assessing its performance in annotating somatic mutations from a published small cell lung cancer study.

Identification of Differentially Expressed Genes in Human Small Cell Lung Carcinoma Using Subtractive Hybridization

  • Ahn Seung-Ju;Choi Jae-Kyoung;Joo Young Mi;Lee Min-A;Choi Pyung-Rak;Lee Yeong-Mi;Kim Myong-Shin;Kim So-Young;Jeon Eun-Hee;Min Byung-In;Kim Chong-Rak
    • Biomedical Science Letters
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    • v.10 no.3
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    • pp.195-202
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    • 2004
  • Lung cancer is a leading cause of cancer death worldwide; however, despite major advances in cancer treatment during the past two decades, the prognostic outcome of lung cancer patients has improved only minimally. This is largely due to the inadequacy of the traditional screening approach of diagnosis in lung cancer, which detects only well­established overt cancers and fails to identify precursor lesions in premalignant conditions of the bronchial tree. In recent years this situation has fundamentally changed with the identification of molecular abnormalities characteristic of premalignant changes; these concern tumour suppressor genes, loss of heterozygosity at crucial sites and activation of oncogenes. Basic knowledge at the molecular level has extremely important clinical implications with regard to early diagnosis, risk assessment and prevention, and therapeutic targets. In this study we used a 'cap-finder' subtractive hybridization method, 'long distance' polymerase chain reaction (PCR), streptavidin magnetic beads mediated subtraction, and spin column chromatography to detect differential expression genes of human small cell lung carcinoma. We have now isolated ninety two genes that expressed differentially in the human small cell lung carcinoma cells and analyzed of 12 clones with sequencing, nine cDNAs include tapasin (NGS-17) mRNA, BC200 alpha scRNA, chromosome 12q24 PAC RPCI3-462E2, protein phosphatase 1 (PPPICA), translocation protein 1 (TLOC1), ribosomal protein S24 (RPS24) mRNA, protein phosphatase (PPEF2), cathepsin Z, MDM2 gene and three novel genes. They may be oncogenesis­related proteins.

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Biological Characteristics of Organic Soil applying Rye (Secale cereal L.) as Green Manure for the Long Term (장기간 호밀을 풋거름작물로 시용한 유기농 토양의 생물학적 특징)

  • Bak, Gye-Ryeong;Lee, Gye-Jun;Kim, Tae-Yeong;Jee, Sam-Nyu;Kim, Chang-Seok;Lee, Hyeong-Bok;Lee, Eun-Kyeong;Song, Jae-Kyeong
    • Korean Journal of Organic Agriculture
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    • v.26 no.3
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    • pp.427-437
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    • 2018
  • In this study, microorganism community characteristics of organic managed soil which applied rye (Secale cereal L.) as green manure for 25 years, were determined. The chemical properties of organic soil showed high level of organic matter and available $P_2O_5$, while the level of exchangeable cation was low. The analysis of dehydrogenase activity and carbon source utilization indicated that the values in on organic soil were significantly higher than those of the control. It suggested that the microorganism community of organic soil had high microorganism activity, compared to the control. In addition, when the 16S rRNA gene-targeted NGS (Next generation sequencing) analysis was conducted to estimate the class of bacterial community, the class level of bacterial taxon composition on organic soil showed higher portion of Sphingobacteriia, Acidobacteriia, Gammaproteobacteria, Solibacteres and Planctomycetia. By base on the results of various reports in which organic managed soil had high portion of Acidobacteriia and Planctomycetia, the characteristic of taxon composition in organic soil, which showed the high percentages of Ktedonobacteria, Sphingobacteriia, Acidobacteriia and Gammaproteobacteria, was resulted from the application of rye as a green manure for the long term. However, further researches were needed because the crop effect was not considered in this study.

Experimental Infection of Different Tomato Genotypes with Tomato mosaic virus Led to a Low Viral Population Heterogeneity in the Capsid Protein Encoding Region

  • Sihelska, Nina;Vozarova, Zuzana;Predajna, Lukas;Soltys, Katarina;Hudcovicova, Martina;Mihalik, Daniel;Kraic, Jan;Mrkvova, Michaela;Kudela, Otakar;Glasa, Miroslav
    • The Plant Pathology Journal
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    • v.33 no.5
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    • pp.508-513
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    • 2017
  • The complete genome sequence of a Slovak SL-1 isolate of Tomato mosaic virus (ToMV) was determined from the next generation sequencing (NGS) data, further confirming a limited sequence divergence in this tobamovirus species. Tomato genotypes Monalbo, Mobaci and Moperou, respectively carrying the susceptible tm-2 allele or the Tm-1 and Tm-2 resistant alleles, were tested for their susceptibility to ToMV SL-1. Although the three tomato genotypes accumulated ToMV SL-1 to similar amounts as judged by semiquantitative DAS-ELISA, they showed variations in the rate of infection and symptomatology. Possible differences in the intra-isolate variability and polymorphism between viral populations propagating in these tomato genotypes were evaluated by analysis of the capsid protein (CP) encoding region. Irrespective of genotype infected, the intra-isolate haplotype structure showed the presence of the same highly dominant CP sequence and the low level of population diversity (0.08-0.19%). Our results suggest that ToMV CP encoding sequence is relatively stable in the viral population during its replication in vivo and provides further demonstration that RNA viruses may show high sequence stability, probably as a result of purifying selection.

Analysis of Pineal Cyst ; Clinical and Radiological Characteristics (송과체 낭종의 임상적, 방사선학적 특성 분석)

  • Kim, Yong-Su;Jung, Shin;Yoon, Sang-Won;Lee, Jung-Kil;Kim, Tae-Sun;Kim, Jae-Hyoo;Kim, Soo-Han;Kang, Sam-Suk;Lee, Je-Hyuk
    • Journal of Korean Neurosurgical Society
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    • v.29 no.7
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    • pp.899-903
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    • 2000
  • Objective : Object : A 7-year retrospective clinical and MRI study was reviewed to evaluate the characteristics of pineal cyst. We evaluated results of clinical and radiologic change of pineal cyst during follow up period. Patients and Methods : From 1992 to 1999, 50 patients with pineal cyst were visited for evaluation. Follow up clinical information was obtained from patients or clinical record through phone conversation. also radiologic findi-ngs of pineal cyst in mid-sagittal MRI were reviewed in detail. Results : Pineal cysts were usually asymptomatic and detected incidentally during evaluation of other diseases. Sagittal MRI is the most useful diagnostic test. The radiologic findings of pineal cyst were isointensity on T1WI comared to CSF and slightly high signal intensity on T2WI. Cyst wall was variably enhanced on Gad-enhanced T1WI. The average size of cyst was $13.9{\times}13.0mm$. The pineal cyst was classified into five groups according to previous report. Long term behavior of these lesions are not apparent, but in our study they showed no specific changes in clinical and radiological aspects during 25 months. Conclusion : Although long-term follow up results are needed, the pineal cyst are considered as normal variants. In our 25 months follow up results, there was no rapid change of cyst both clinically and radiologically. Therefore, frequent radiologic evaluation seems unnecessary except type III pineal cyst.

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A Case Study of Bottom Liner Construction Using Composite Liner Technology in a Solid Waste Landfill (복합차수층 조성기술을 이용한 폐기물매립지 바닥차수층 시공사례)

  • Lee, Kyu-Jung;Lee, Nam-Hoon;Park, Soo-Young;Jeon, Won-Pyo;Lee, Chae-Young
    • Journal of the Korea Organic Resources Recycling Association
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    • v.15 no.3
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    • pp.90-96
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    • 2007
  • Modern waste management units, so-called "landfills" protect human health and environment from hazardous leachate and gas. Accordingly, it must be constructed with a bottom liner system that includes a gas collection layer. Leachate is the contaminated liquid that drains from the waste material pollutes ground water. For this reason. bottom liner system must have durability and low hydraulic conductivity (in case of compacted clay liner, no more than $1{\times}10^{-7}cm/sec$ ). P county in Kangwon province constructed a solid waste landfill with bottom liner system. In this study. it is mainly introduced that the test results on construction and quality control of bottom liner system by "Multiple composite liner construction technology", which is selected for bottom liner system in P solid waste landfill.

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Assessment of the Dynamics of Microbial Community Associated with Tetraselmis suecica Culture under Different LED Lights Using Next-Generation Sequencing

  • Yang, Su-Jeong;Kim, Hyun-Woo;Choi, Seok-Gwan;Chung, Sangdeok;Oh, Seok Jin;Borkar, Shweta;Kim, Hak Jun
    • Journal of Microbiology and Biotechnology
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    • v.29 no.12
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    • pp.1957-1968
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    • 2019
  • Tetraselmis is a green algal genus, some of whose species are important in aquaculture as well as biotechnology. In algal culture, fluorescent lamps, traditional light source for culturing algae, are now being replaced by a cost-effective light-emitting diodes (LEDs). In this study, we investigated the effect of LED light of different wavelengths (white, red, yellow, and blue) on the growth of Tetraselmis suecica and its associated microbial community structures using the next-generation sequencing (NGS). The fastest growth rate of T. suecica was shown in the red light, whereas the slowest was in yellow. The highest OTUs (3426) were identified on day 0, whereas the lowest ones (308) were found on day 15 under red light. The top 100 OTUs associated with day 0 and day 5 cultures of T. suecica under the red and yellow LED were compared. Only 26 OTUs were commonly identified among four samples. The highest numbers of unique OTUs were identified at day 0, indicating the high degree of initial microbial diversity of the T. suecica inoculum. The red light-unique OTUs occupied 34.98%, whereas the yellow-specific OTUs accounted for only 2.2%. This result suggested a higher degree of interaction in T. suecica culture under the red light, where stronger photosynthesis occurs. Apparently, the microbial community associated with T. suecica related to the oxygen produced by algal photosynthesis. This result may expand our knowledge about the algae-bacteria consortia, which would be useful for various biotechnological applications including wastewater treatment, bioremediation, and sustainable aquaculture.

Application of Solanum lycopersicum Glucose-6-phosphate Dehydrogenase to NADPH-generating System for Cytochrome P450 Reactions

  • Park, Chan Mi;Jeong, Heon;Ma, Sang Hoon;Kim, Hyun Min;Joung, Young Hee;Yun, Chul-Ho
    • Microbiology and Biotechnology Letters
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    • v.47 no.4
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    • pp.536-545
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    • 2019
  • Cytochrome P450 (P450 or CYP) is involved in the metabolism of endogenous and exogenous compounds in most organisms. P450s have great potential as biocatalysts in the pharmaceutical and fine chemical industries because they catalyze diverse oxidative reactions using a wide range of substrates. The high-cost nicotinamide cofactor, NADPH, is essential for P450 reactions. Glucose-6-phosphate dehydrogenase (G6PDH) has been commonly used in NADPH-generating systems (NGSs) to provide NADPH for P450 reactions. Currently, only two G6PDHs from Leuconostoc mesenteroides and Saccharomyces cerevisiae can be obtained commercially. To supply high-cost G6PDH cost-effectively, we cloned the cytosolic G6PDH gene of Solanum lycopersicum (tomato) with 6xHis tag, expressed it in Escherichia coli, and purified the recombinant G6PDH (His-G6PDH) using affinity chromatography. In addition, enzymatic properties of His-G6PDH were investigated, and the His-G6PDH-coupled NGS was optimized for P450 reactions. His-G6PDH supported CYP102A1-catalyzed hydroxylation of omeprazole and testosterone by NADPH generation. This result suggests that tomato His-G6PDH could be a cost-effective enzyme source for NGSs for P450-catalyzed reactions as well as other NADPH-requiring reactions.

Massive Parallel Sequencing for Diagnostic Genetic Testing of BRCA Genes - a Single Center Experience

  • Ermolenko, Natalya A;Boyarskikh, Uljana A;Kechin, Andrey A;Mazitova, Alexandra M;Khrapov, Evgeny A;Petrova, Valentina D;Lazarev, Alexandr F;Kushlinskii, Nikolay E;Filipenko, Maxim L
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.17
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    • pp.7935-7941
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    • 2015
  • The aim of this study was to implement massive parallel sequencing (MPS) technology in clinical genetics testing. We developed and tested an amplicon-based method for resequencing the BRCA1 and BRCA2 genes on an Illumina MiSeq to identify disease-causing mutations in patients with hereditary breast or ovarian cancer (HBOC). The coding regions of BRCA1 and BRCA2 were resequenced in 96 HBOC patient DNA samples obtained from different sample types: peripheral blood leukocytes, whole blood drops dried on paper, and buccal wash epithelia. A total of 16 random DNA samples were characterized using standard Sanger sequencing and applied to optimize the variant calling process and evaluate the accuracy of the MPS-method. The best bioinformatics workflow included the filtration of variants using GATK with the following cut-offs: variant frequency >14%, coverage ($>25{\times}$) and presence in both the forward and reverse reads. The MPS method had 100% sensitivity and 94.4% specificity. Similar accuracy levels were achieved for DNA obtained from the different sample types. The workflow presented herein requires low amounts of DNA samples (170 ng) and is cost-effective due to the elimination of DNA and PCR product normalization steps.