• Title/Summary/Keyword: High-throughput Sequencing

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Effect of Soil Properties and Soil Bacterial Community on Early Growth Characteristics of Wild-simulated Ginseng (Panax ginseng C. A. Meyer) in Coniferous and Mixed Forest (침엽수림과 혼효림에서 토양특성과 토양세균 군집이 산양삼 초기 생육특성에 미치는 영향)

  • Kim, Ki Yoon;Kim, Hyun Jun;Um, Yurry;Jeon, Kwon Seok
    • Korean Journal of Medicinal Crop Science
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    • v.28 no.3
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    • pp.183-194
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    • 2020
  • Background: This study investigated the effect of soil properties and soil bacterial community on early growth characteristics of wild-simulated ginseng (Panax ginseng C. A. Meyer) in coniferous and mixed forest experimental fields. Methods and Results: The soil bacterial community was analyzed using a high throughput sequencing technique (Illumina MiSeq sequencing). The relationship between the soil bacterial community, soil properties, and growth characteristics of wild-simulated ginseng were analyzed using principal coordinate analysis (PCoA) and the Pearson's correlation analysis. Soil properties and soil bacterial community showed significant difference with forest physiognomy. Results of Pearson's correlation analysis and PCoA showed that the soil properties (soil pH, organic matter, total nitrogen, and cation exchange capacity) and soil bacterial community had significant correlation with tree species ratio and early growth characteristics of wild-simulated ginseng. Conclusions: This study clearly demonstrated the effect of soil properties and soil bacterial community on early growth characteristics of wild-simulated ginseng in coniferous and mixed forest. Moreover, these results will help in the selection of suitable cultivation sites for wild-simulated ginseng.

Identification of causative mutations in patients with Leigh syndrome and MERRF by mitochondrial DNA-targeted next-generation sequencing

  • Hong, Hyun Dae;Kim, Eunja;Nam, Soo Hyun;Yoo, Da Hye;Suh, Bum Chun;Choi, Byung-Ok;Chung, Ki Wha
    • Journal of Genetic Medicine
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    • v.12 no.2
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    • pp.109-117
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    • 2015
  • Purpose: Mitochondrial diseases are clinically and genetically heterogeneous disorders, which make their exact diagnosis and classification difficult. The purpose of this study was to identify pathogenic mitochondrial DNA (mtDNA) mutations in 2 Korean families with myoclonic epilepsy with ragged-red fibers (MERRF) and Leigh syndrome, respectively. Materials and Methods: Whole mtDNAs were sequenced by the method of mtDNA-targeted next-generation sequencing (NGS). Results: Two causative mtDNA mutations were identified from the NGS data. An m.8344A>G mutation in the tRNA-Lys gene (MT-TK) was detected in a MERRF patient (family ID: MT132), and an m.9176T>C (p.Leu217Pro) mutation in the mitochondrial ATP6 gene (MT-ATP6) was detected in a Leigh syndrome patient (family ID: MT130). Both mutations, which have been reported several times before in affected individuals, were not found in the control samples. Conclusion: This study suggests that mtDNA-targeted NGS will be helpful for the molecular diagnosis of genetically heterogeneous mitochondrial diseases with complex phenotypes.

Effects of husbandry systems and Chinese indigenous chicken strain on cecum microbial diversity

  • Dong, Xiuxue;Hu, Bing;Wan, Wenlong;Gong, Yanzhang;Feng, Yanping
    • Asian-Australasian Journal of Animal Sciences
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    • v.33 no.10
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    • pp.1610-1616
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    • 2020
  • Objective: This study was to evaluate the effect of husbandry systems and strains on cecum microbial diversity of Jingyang chickens under the same dietary conditions. Methods: A total of 320 laying hens (body weight, 1.70±0.15 kg; 47 weeks old) were randomly allocated to one of the four treatments: i) Silver-feathered hens in enrichment cages (SEC) with an individual cage (70×60×75 cm), ii) Silver-feathered hens in free range (SFR) with the stocking density of 1.5 chickens per ten square meters, iii) Gold-feathered hens in enrichment cages (GEC), iv) Gold-feathered hens in free range (GFR). The experiment lasted 8 weeks and the cecum fecal samples were collected for 16S rDNA high throughput sequencing at the end of experiment. Results: i) The core microbiota was composed of Bacteroidetes (49% to 60%), Firmicutes (21% to 32%) and Proteobacteria (2% to 4%) at the phylum level. ii) The core bacteria were Bacteroides (26% to 31%), Rikenellaceae (9% to 16%), Parabacteroides (2% to 5%) and Lachnoclostridium (2% to 6%) at the genus level. iii) The indexes of operational taxonomic unit, Shannon, Simpson and observed species were all higher in SFR group than in SEC group while in GEC group than in GFR group, with SFR group showing the greatest diversity of cecum microorganisms among the four groups. iv) The clustering result was consistent with the strain classification, with a similar composition of cecum bacteria in the two strains of laying hens. Conclusion: The core microbiota were not altered by husbandry systems or strains. The free-range system increased the diversity of cecal microbes only for silver feathered hens. However, the cecum microbial composition was similar in two strain treatments under the same dietary conditions.

TGF-β Signaling and miRNAs Targeting for BMP7 in the Spleen of Two Necrotic Enteritis-Afflicted Chicken Lines

  • Truong, Anh Duc;Hong, Yeojin;Lee, Janggeun;Lee, Kyungbaek;Lillehoj, Hyun S.;Hong, Yeong Ho
    • Korean Journal of Poultry Science
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    • v.44 no.3
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    • pp.211-223
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    • 2017
  • Transforming growth factor beta ($TGF-{\beta}$) signaling pathways are involved in the regulation of proliferation, differentiation, immunity, survival, and apoptosis of many cells. The aim of this study was to investigate the differential expression of $TGF-{\beta}$-related genes, and their interactions and regulators in the spleen of two genetically disparate chicken lines (Marek's disease resistant line 6.3 and Marek's disease-susceptible line 7.2) induced with necrotic enteritis (NE) by Eimeria maxima and Clostridium perfringens infection. By using high-throughput RNA-sequencing, we investigated 76 $TGF-{\beta}$-related genes that were significantly and differentially expressed in the spleens of the chickens. Approximately 20 $TGF-{\beta}$ pathway genes were further verified by qRT-PCR, and the results were consistent with our RNA sequencing data. All 76 identified genes were analyzed through Gene Ontology and mapped onto the KEGG chicken $TGF-{\beta}$ pathway. Our results demonstrated that several key genes, including $TGF-{\beta}$1-3, bone morphogenetic proteins (BMP)1-7, inhibitor of differentiation (ID) proteins ID1-3, SMAD1-9, and Jun, showed a markedly differential expression between the two chicken lines, relative to their respective controls. We then further predicted 24 known miRNAs that targeted BMP7 mRNA from 139 known miRNAs in the two chicken lines. Among these, six miRNAs were measured by qRT-PCR. In conclusion, this study is the first to analyze most of the genes, interactions, and regulators of the $TGF-{\beta}$ pathway in the innate immune responses of NE afflicted chickens.

Comprehensive proteome analysis using quantitative proteomic technologies

  • Kamal, Abu Hena Mostafa;Choi, Jong-Soon;Cho, Yong-Gu;Kim, Hong-Sig;Song, Beom-Heon;Lee, Chul-Won;Woo, Sun-Hee
    • Journal of Plant Biotechnology
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    • v.37 no.2
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    • pp.196-204
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    • 2010
  • With the completion of genome sequencing of several organisms, attention has been focused to determine the function and functional network of proteins by proteome analysis. The recent techniques of proteomics have been advanced quickly so that the high-throughput and systematic analyses of cellular proteins are enabled in combination with bioinformatics tools. Furthermore, the development of proteomic techniques helps to elucidate the functions of proteins under stress or diseased condition, resulting in the discovery of biomarkers responsible for the biological stimuli. Ultimate goal of proteomics orients toward the entire proteome of life, subcellular localization, biochemical activities, and their regulation. Comprehensive analysis strategies of proteomics can be classified as three categories: (i) protein separation by 2-dimensional gel electrophoresis (2-DE) or liquid chromatography (LC), (ii) protein identification by either Edman sequencing or mass spectrometry (MS), and (iii) quanitation of proteome. Currently MS-based proteomics turns shiftly from qualitative proteome analysis by 2-DE or 2D-LC coupled with off-line matrix assisted laser desorption ionization (MALDI) and on-line electrospray ionization (ESI) MS, respectively, to quantitative proteome analysis. Some new techniques which include top-down mass spectrometry and tandem affinity purification have emerged. The in vitro quantitative proteomic techniques include differential gel electrophoresis with fluorescence dyes, protein-labeling tagging with isotope-coded affinity tag, and peptide-labeling tagging with isobaric tags for relative and absolute quantitation. In addition, stable isotope labeled amino acid can be in vivo labeled into live culture cells through metabolic incorporation. MS-based proteomics extends to detect the phosphopeptide mapping of biologically crucial protein known as one of post-translational modification. These complementary proteomic techniques contribute to not only the understanding of basic biological function but also the application to the applied sciences for industry.

EST Knowledge Integrated Systems (EKIS): An Integrated Database of EST Information for Research Application

  • Kim, Dae-Won;Jung, Tae-Sung;Choi, Young-Sang;Nam, Seong-Hyeuk;Kwon, Hyuk-Ryul;Kim, Dong-Wook;Choi, Han-Suk;Choi, Sang-Heang;Park, Hong-Seog
    • Genomics & Informatics
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    • v.7 no.1
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    • pp.38-40
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    • 2009
  • The EST Knowledge Integrated System, EKIS (http://ekis.kribb.re.kr), was established as a part of Korea's Ministry of Education, Science and Technology initiative for genome sequencing and application research of the biological model organisms (GEAR) project. The goals of the EKIS are to collect EST information from GEAR projects and make an integrated database to provide transcriptomic and metabolomic information for biological scientists. The EKIS constitutes five independent categories and several retrieval systems in each category for incorporating massive EST data from high-throughput sequencing of 65 different species. Through the EKIS database, scientists can freely access information including BLAST functional annotation as well as Genechip and pathway information for KEGG. By integrating complex data into a framework of existing EST knowledge information, the EKIS provides new insights into specialized metabolic pathway information for an applied industrial material.

Anticancer Activity of the Antimicrobial Peptide Scolopendrasin VII Derived from the Centipede, Scolopendra subspinipes mutilans

  • Lee, Joon Ha;Kim, In-Woo;Kim, Sang-Hee;Kim, Mi-Ae;Yun, Eun-Young;Nam, Sung-Hee;Ahn, Mi-Young;Kang, Dongchul;Hwang, Jae Sam
    • Journal of Microbiology and Biotechnology
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    • v.25 no.8
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    • pp.1275-1280
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    • 2015
  • Previously, we performed de novo RNA sequencing of Scolopendra subspinipes mutilans using high-throughput sequencing technology and identified several antimicrobial peptide candidates. Among them, a cationic antimicrobial peptide, scolopendrasin VII, was selected based on its physicochemical properties, such as length, charge, and isoelectric point. Here, we assessed the anticancer activities of scolopendrasin VII against U937 and Jurkat leukemia cell lines. The results showed that scolopendrasin VII decreased the viability of the leukemia cells in MTS assays. Furthermore, flow cytometric analysis and acridine orange/ethidium bromide staining revealed that scolopendrasin VII induced necrosis in the leukemia cells. Scolopendrasin VII-induced necrosis was mediated by specific interaction with phosphatidylserine, which is enriched in the membrane of cancer cells. Taken together, these data indicated that scolopendrasin VII induced necrotic cell death in leukemia cells, probably through interaction with phosphatidylserine. The results provide a useful anticancer peptide candidate and an efficient strategy for new anticancer peptide development.

Characterization of the bacterial microbiota across the different intestinal segments of the Qinghai semi-fine wool sheep on the Qinghai-Tibetan Plateau

  • Wang, Xungang;Hu, Linyong;Liu, Hongjin;Xu, Tianwei;Zhao, Na;Zhang, Xiaoling;Geng, Yuanyue;Kang, Shengping;Xu, Shixiao
    • Animal Bioscience
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    • v.34 no.12
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    • pp.1921-1929
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    • 2021
  • Objective: The intestinal microbiota enhances nutrient absorption in the host and thus promotes heath. Qinghai semi-fine wool sheep is an important livestock raised in the Qinghai-Tibetan Plateau; however, little is known about the bacterial microbiota of its intestinal tract. The aim of this study was to detect the microbial characterization in the intestinal tract of the Qinghai semi-fine wool sheep. Methods: The bacterial profiles of the six different intestinal segments (duodenum, jejunum, ileum, cecum, colon and rectum) of Qinghai semi-fine wool sheep were studied using 16S rRNA V3-V4 hypervariable amplicon sequencing. Results: A total of 2,623,323 effective sequences were obtained, and 441 OTUs shared all six intestinal segments. The bacterial diversity was significantly different among the different intestinal segments, and the large intestine exhibited higher bacterial diversity than the small intestine. Firmicutes, Bacteroidetes, and Patescibacteria were the dominant phyla in these bacterial communities. Additionally, at the genus level, Prevotella_1, Candidatus_Saccharimonas, and Ruminococcaceae_UCG-005 were the most predominant genus in duodenal segment, jejunal and ileal segments, and cecal, colonic, and rectal segments, respectively. We predicted that the microbial functions and the relative abundance of the genes involved in carbohydrate metabolism were overrepresented in the intestinal segments of Qinghai semi-fine wool sheep. Conclusion: The bacterial communities and functions differed among different intestinal segments. Our study is the first to provide insights into the composition and biological functions of the intestinal microbiota of Qinghai semi-fine wool sheep. Our results also provide useful information for the nutritional regulation and production development in Qinghai semi-fine wool sheep.

Comparative analysis of commonly used peak calling programs for ChIP-Seq analysis

  • Jeon, Hyeongrin;Lee, Hyunji;Kang, Byunghee;Jang, Insoon;Roh, Tae-Young
    • Genomics & Informatics
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    • v.18 no.4
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    • pp.42.1-42.9
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    • 2020
  • Chromatin immunoprecipitation coupled with high-throughput DNA sequencing (ChIP-Seq) is a powerful technology to profile the location of proteins of interest on a whole-genome scale. To identify the enrichment location of proteins, many programs and algorithms have been proposed. However, none of the commonly used peak calling programs could accurately explain the binding features of target proteins detected by ChIP-Seq. Here, publicly available data on 12 histone modifications, including H3K4ac/me1/me2/me3, H3K9ac/me3, H3K27ac/me3, H3K36me3, H3K56ac, and H3K79me1/me2, generated from a human embryonic stem cell line (H1), were profiled with five peak callers (CisGenome, MACS1, MACS2, PeakSeq, and SISSRs). The performance of the peak calling programs was compared in terms of reproducibility between replicates, examination of enriched regions to variable sequencing depths, the specificity-to-noise signal, and sensitivity of peak prediction. There were no major differences among peak callers when analyzing point source histone modifications. The peak calling results from histone modifications with low fidelity, such as H3K4ac, H3K56ac, and H3K79me1/me2, showed low performance in all parameters, which indicates that their peak positions might not be located accurately. Our comparative results could provide a helpful guide to choose a suitable peak calling program for specific histone modifications.

Lower Airway Microbiota and Lung Cancer

  • Sanchez-Hellin, Victoria;Galiana, Antonio;Zamora-Molina, Lucia;Soler-Sempere, Maria J.;Grau-Delgado, Justo;Barbera, Victor M.;Padilla-Navas, Isabel;Garcia-Pachon, Eduardo
    • Microbiology and Biotechnology Letters
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    • v.47 no.3
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    • pp.441-448
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    • 2019
  • This study was aimed at identifying the lower airway microbiota in patients with lung cancer (LC) using protected brush sampling. We enrolled 37 patients undergoing diagnostic bronchoscopy for suspected LC, 26 with LC and 11 with benign diseases. Protected brush specimens were obtained from the contralateral lung and the side of the tumor; these specimens were analyzed by 16S rRNA-based-next-generation sequencing. The results indicated that the biodiversity was not different between groups, and there were no significant differences between the proportion of microorganisms in the tumor and in the contralateral side of patients with LC. In patients with LC, there was a higher abundance of several microorganisms including Capnocytophaga, Haemophilus, Enterococcus, and Streptococcus; whereas, in individuals without LC, Bacteroides, Lactobacillus, or Methylobacterium were more abundant. Malignancy could be determined with an accuracy of 70% by isolating Enterococcus, Capnocytophaga, or Actinomyces. Microbispora indicated benignity with a sensitivity of 55%, specificity of 88%, and accuracy of 78%. Lower airway microbiota in patients with LC is fairly similar in both the tumor and contralateral sites. Endobronchial microbiota is different in patients with and without LC, and these differences may have a potential clinical value as diagnostic or prognostic biomarkers.