• Title/Summary/Keyword: NEXT GENERATION SEQUENCING

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Characterization of the Biodiversity of the Spoilage Microbiota in Chicken Meat Using Next Generation Sequencing and Culture Dependent Approach

  • Lee, Hee Soo;Kwon, Mirae;Heo, Sunhak;Kim, Min Gon;Kim, Geun-Bae
    • Food Science of Animal Resources
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    • v.37 no.4
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    • pp.535-541
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    • 2017
  • This study investigated the psychrotrophic bacteria isolated from chicken meat to characterize their microbial composition during refrigerated storage. The bacterial community was identified by the Illumina MiSeq method based on bacterial DNA extracted from spoiled chicken meat. Molecular identification of the isolated psychrotrophic bacteria was carried out using 16S rDNA sequencing and their putrefactive potential was investigated by the growth at low temperature as well as their proteolytic activities in chicken meat. From the Illumina sequencing, a total of 187,671 reads were obtained from 12 chicken samples. Regardless of the type of chicken meat (i.e., whole meat and chicken breast) and storage temperatures ($4^{\circ}C$ and $10^{\circ}C$), Pseudomonas weihenstephanensis and Pseudomonas congelans were the most prominent bacterial species. Serratia spp. and Acinetobacter spp. were prominent in chicken breast and whole chicken meat, respectively. The 118 isolated strains of psychrotrophic bacteria comprised Pseudomonas spp. (58.48%), Serratia spp. (10.17%), and Morganella spp. (6.78%). All isolates grew well at $10^{\circ}C$ and they induced different proteolytic activities depending on the species and strains. Parallel analysis of the next generation sequencing and culture dependent approach provides in-depth information on the biodiversity of the spoilage microbiota in chicken meat. Further study is needed to develop better preservation methods against these spoilage bacteria.

Application of Amplicon Pyrosequencing in Soil Microbial Ecology (토양미생물 생태 연구를 위한 증폭 파이로시퀀싱 기법의 응용)

  • Ahn, Jae-Hyung;Kim, Byung-Yong;Kim, Dae-Hoon;Song, Jaekyeong;Weon, Hang-Yeon
    • Korean Journal of Soil Science and Fertilizer
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    • v.45 no.6
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    • pp.1073-1085
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    • 2012
  • Soil microbial communities are immensely diverse and complex with respect to species richness and community size. These communities play essential roles in agricultural soil because they are responsible for most of the nutrient cycles in the soil and influence the plant diversity and productivity. However, the majority of these microbes remain uncharacterized because of poor culturability. Next-generation sequencing techniques have revolutionized many areas of biology by providing cheaper and faster alternatives to Sanger sequencing. Among them, amplicon pyrosequencing is a powerful tool developed by 454 Life Sciences for assessing the diversity of complex microbial communities by sequencing PCR products or amplicons. This review summarizes the current opinions in amplicon sequencing of soil microbial communities, and provides practical guidance and advice on sequence quality control, aligning, clustering, OTU- and taxon-based analysis. The last section of this article includes a few representative studies conducted using amplicon pyrosequencing.

Caution and Curation for Complete Mitochondrial Genome from Next-Generation Sequencing: A Case Study from Dermatobranchus otome (Gastropoda, Nudibranchia)

  • Do, Thinh Dinh;Choi, Yisoo;Jung, Dae-Wui;Kim, Chang-Bae
    • Animal Systematics, Evolution and Diversity
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    • v.36 no.4
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    • pp.336-346
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    • 2020
  • Mitochondrial genome is an important molecule for systematic and evolutionary studies in metazoans. The development of next-generation sequencing (NGS) technique has rapidly increased the number of mitogenome sequences. The process of generating mitochondrial genome based on NGS includes different steps, from DNA preparation, sequencing, assembly, and annotation. Despite the effort to improve sequencing, assembly, and annotation methods of mitogenome, the low quality and/or quantity sequence in the final map can still be generated through the work. Therefore, it is necessary to check and curate mitochondrial genome sequence after annotation for proofreading and feedback. In this study, we introduce the pipeline for sequencing and curation for mitogenome based on NGS. For this purpose, two mitogenome sequences of Dermatobranchus otome were sequenced by Illumina Miseq system with different amount of raw read data. Generated reads were targeted for assembly and annotation with commonly used programs. As abnormal repeat regions present in the mitogenomes after annotation, primers covering these regions were designed and conventional PCR followed by Sanger sequencing were performed to curate the mitogenome sequences. The obtained sequences were used to replace the abnormal region. Following the replacement, each mitochondrial genome was compared with the other as well as the sequences of close species available on the Genbank for confirmation. After curation, two mitogenomes of D. otome showed a typically circular molecule with 14,559 bp in size and contained 13 protein-coding genes, 22 tRNA genes, two rRNA genes. The phylogenetic tree revealed a close relationship between D. otome and Tritonia diomea. The finding of this study indicated the importance of caution and curation for the generation of mitogenome from NGS.

Challenges of Genome Wide Sequencing Technologies in Prenatal Medicine (산전 진단에서의 염기 서열 분석 방법의 의의)

  • Kang, Ji-Un
    • The Journal of the Korea Contents Association
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    • v.22 no.2
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    • pp.762-769
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    • 2022
  • Genetic testing in prenatal diagnosis is a precious tool providing valuable information in clinical management and parental decision-making. For the last year, cytogenetic testing methods, such as G-banding karyotype analysis, fluorescent in situ hybridization, chromosomal microarray, and gene panels have evolved to become part of routine laboratory testing. However, the limitations of each of these methods demonstrate the need for a revolutionary technology that can alleviate the need for multiple technologies. The recent introduction of new genomic technologies based on next-generation sequencing has changed the current practice of prenatal testing. The promise of these innovations lies in the fast and cost-effective generation of genome-scale sequence data with exquisite resolution and accuracy for prenatal diagnosis. Here, we review the current state of sequencing-based pediatric diagnostics, associated challenges, as well as future prospects.

Next Generation Sequencing and Bioinformatics (차세대 염기서열 분석기법과 생물정보학)

  • Kim, Ki-Bong
    • Journal of Life Science
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    • v.25 no.3
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    • pp.357-367
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    • 2015
  • With the ongoing development of next-generation sequencing (NGS) platforms and advancements in the latest bioinformatics tools at an unprecedented pace, the ultimate goal of sequencing the human genome for less than $1,000 can be feasible in the near future. The rapid technological advances in NGS have brought about increasing demands for statistical methods and bioinformatics tools for the analysis and management of NGS data. Even in the early stages of the commercial availability of NGS platforms, a large number of applications or tools already existed for analyzing, interpreting, and visualizing NGS data. However, the availability of this plethora of NGS data presents a significant challenge for storage, analyses, and data management. Intrinsically, the analysis of NGS data includes the alignment of sequence reads to a reference, base-calling, and/or polymorphism detection, de novo assembly from paired or unpaired reads, structural variant detection, and genome browsing. While the NGS technologies have allowed a massive increase in available raw sequence data, a number of new informatics challenges and difficulties must be addressed to improve the current state and fulfill the promise of genome research. This review aims to provide an overview of major NGS technologies and bioinformatics tools for NGS data analyses.

Efficiency to Discovery Transgenic Loci in GM Rice Using Next Generation Sequencing Whole Genome Re-sequencing

  • Park, Doori;Kim, Dongin;Jang, Green;Lim, Jongsung;Shin, Yun-Ji;Kim, Jina;Seo, Mi-Seong;Park, Su-Hyun;Kim, Ju-Kon;Kwon, Tae-Ho;Choi, Ik-Young
    • Genomics & Informatics
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    • v.13 no.3
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    • pp.81-85
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    • 2015
  • Molecular characterization technology in genetically modified organisms, in addition to how transgenic biotechnologies are developed now require full transparency to assess the risk to living modified and non-modified organisms. Next generation sequencing (NGS) methodology is suggested as an effective means in genome characterization and detection of transgenic insertion locations. In the present study, we applied NGS to insert transgenic loci, specifically the epidermal growth factor (EGF) in genetically modified rice cells. A total of 29.3 Gb (${\sim}72{\times}coverage$) was sequenced with a $2{\times}150bp$ paired end method by Illumina HiSeq2500, which was consecutively mapped to the rice genome and T-vector sequence. The compatible pairs of reads were successfully mapped to 10 loci on the rice chromosome and vector sequences were validated to the insertion location by polymerase chain reaction (PCR) amplification. The EGF transgenic site was confirmed only on chromosome 4 by PCR. Results of this study demonstrated the success of NGS data to characterize the rice genome. Bioinformatics analyses must be developed in association with NGS data to identify highly accurate transgenic sites.

Bacterial Community and Diversity from the Watermelon Cultivated Soils through Next Generation Sequencing Approach

  • Adhikari, Mahesh;Kim, Sang Woo;Kim, Hyun Seung;Kim, Ki Young;Park, Hyo Bin;Kim, Ki Jung;Lee, Youn Su
    • The Plant Pathology Journal
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    • v.37 no.6
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    • pp.521-532
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    • 2021
  • Knowledge and better understanding of functions of the microbial community are pivotal for crop management. This study was conducted to study bacterial structures including Acidovorax species community structures and diversity from the watermelon cultivated soils in different regions of South Korea. In this study, soil samples were collected from watermelon cultivation areas from various places of South Korea and microbiome analysis was performed to analyze bacterial communities including Acidovorax species community. Next generation sequencing (NGS) was performed by extracting genomic DNA from 92 soil samples from 8 different provinces using a fast genomic DNA extraction kit. NGS data analysis results revealed that, total, 39,367 operational taxonomic unit (OTU), were obtained. NGS data results revealed that, most dominant phylum in all the soil samples was Proteobacteria (37.3%). In addition, most abundant genus was Acidobacterium (1.8%) in all the samples. In order to analyze species diversity among the collected soil samples, OTUs, community diversity, and Shannon index were measured. Shannon (9.297) and inverse Simpson (0.996) were found to have the highest diversity scores in the greenhouse soil sample of Gyeonggi-do province (GG4). Results from NGS sequencing suggest that, most of the soil samples consists of similar trend of bacterial community and diversity. Environmental factors play a key role in shaping the bacterial community and diversity. In order to address this statement, further correlation analysis between soil physical and chemical parameters with dominant bacterial community will be carried out to observe their interactions.

Analytical Tools and Databases for Metagenomics in the Next-Generation Sequencing Era

  • Kim, Mincheol;Lee, Ki-Hyun;Yoon, Seok-Whan;Kim, Bong-Soo;Chun, Jongsik;Yi, Hana
    • Genomics & Informatics
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    • v.11 no.3
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    • pp.102-113
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    • 2013
  • Metagenomics has become one of the indispensable tools in microbial ecology for the last few decades, and a new revolution in metagenomic studies is now about to begin, with the help of recent advances of sequencing techniques. The massive data production and substantial cost reduction in next-generation sequencing have led to the rapid growth of metagenomic research both quantitatively and qualitatively. It is evident that metagenomics will be a standard tool for studying the diversity and function of microbes in the near future, as fingerprinting methods did previously. As the speed of data accumulation is accelerating, bioinformatic tools and associated databases for handling those datasets have become more urgent and necessary. To facilitate the bioinformatics analysis of metagenomic data, we review some recent tools and databases that are used widely in this field and give insights into the current challenges and future of metagenomics from a bioinformatics perspective.