Next Generation Sequencing (NGS), A Key Tool to open the Personalized Medicine Era

( Sun-il Kwon ) 대한임상검사과학회 2012 대한임상검사과학회지(KJCLS) Vol.44 No.4

Next-Generation Sequencing (NGS) is a term that means post-Sanger sequencing methods with highthroughput sequencing technologies. NGS parallelizes the sequencing process, producing thousands or millions of sequences at once. The latest NGS technologies use even single DNA molecule as a template and measures the DNA sequence directly via measuring electronic signals from the extension or degradation of DNA. NGS is making big impacts on biomedical research, molecular diagnosis and personalized medicine. The hospitals are rapidly adopting the use of NGS to help to patients understand treatment with sequencing data. As NGS equipments are getting smaller and affordable, many hospitals are in the process of setting up NGS platforms. In this review, the progress of NGS technology development and action mechanisms of representative NGS equipments of each generation were discussed. The key technological advances in the commercialized platforms were presented. As NGS platforms are a great concern in the healthcare area, the latest trend in the use of NGS and the prospect of NGS in the future in diagnosis and personalized medicine were also discussed.

NGS_SNPAnalyzer: a desktop software supporting genome projects by identifying and visualizing sequence variations from next-generation sequencing data

Dong‑Jun Lee,Taesoo Kwon,Chang‑Kug Kim,Young‑Joo Seol,Dong‑Suk Park,Tae‑Ho Lee,Byung‑Ohg Ahn 한국유전학회 2020 Genes & Genomics Vol.42 No.11

Background Sequence variations such as single nucleotide polymorphisms are markers for genetic diseases and breeding. Therefore, identifying sequence variations is one of the main objectives of several genome projects. Although most genomeproject consortiums provide standard operation procedures for sequence variation detection methods, there may be differencesin the results because of human selection or error. Objective To standardize the procedure for sequence variation detection and help researchers who are not formally trainedin bioinformatics, we developed the NGS_SNPAnalyzer, a desktop software and fully automated graphical pipeline. Methods The NGS_SNPAnalyzer is implemented using JavaFX (version 1.8); therefore, it is not limited to any operatingsystem (OS). The tools employed in the NGS_SNPAnalyzer were compiled on Microsoft Windows (version 7, 10) andUbuntu Linux (version 16.04, 17.0.4). Results The NGS_SNPAnalyzer not only includes the functionalities for variant calling and annotation but also providesquality control, mapping, and filtering details to support all procedures from next-generation sequencing (NGS) data to variantvisualization. It can be executed using pre-set pipelines and options and customized via user-specified options. Additionally,the NGS_SNPAnalyzer provides a user-friendly graphical interface and can be installed on any OS that supports JAVA. Conclusions Although there are several pipelines and visualization tools available for NGS data analysis, we developedthe NGS_SNPAnalyzer to provide the user with an easy-to-use interface. The benchmark test results indicate that theNGS_SNPAnayzer achieves better performance than other open source tools.

Genetic tests by next-generation sequencing in children with developmental delay and/or intellectual disability

Han, Ji Yoon,Lee, In Goo The Korean Pediatric Society 2020 Clinical and Experimental Pediatrics (CEP) Vol.63 No.6

Developments in next-generation sequencing (NGS) techogies have assisted in clarifying the diagnosis and treatment of developmental delay/intellectual disability (DD/ID) via molecular genetic testing. Advances in DNA sequencing technology have not only allowed the evolution of targeted panels but also, and more currently enabled genome-wide analyses to progress from research era to clinical practice. Broad acceptance of accuracy-guided targeted gene panel, whole-exome sequencing (WES), and whole-genome sequencing (WGS) for DD/ID need prospective analyses of the increasing cost-effectiveness versus conventional genetic testing. Choosing the appropriate sequencing method requires individual planning. Data are required to guide best-practice recommendations for genomic testing, regarding various clinical phenotypes in an etiologic approach. Targeted panel testing may be recommended as a firsttier testing approach for children with DD/ID. Family-based trio testing by WES/WGS can be used as a second test for DD/ID in undiagnosed children who previously tested negative on a targeted panel. The role of NGS in molecular diagnostics, treatment, prediction of prognosis will continue to increase further in the coming years. Given the rapid pace of changes in the past 10 years, all medical providers should be aware of the changes in the transformative genetics field.

Next-Generation Sequencing Based Transposon Display to Detect High-Throughput Insertion Polymorphism Markers in Brassica

( Sampath Perumal ),( Nomar Espinosa Waminal ),( Jonghoon Lee ),( Nur Kholilatul Izzah ),( Mina Jin ),( Beom-soon Choi ),( Tae-jin Yang ) 한국육종학회 2016 Plant Breeding and Biotechnology Vol.4 No.3

Miniature transposable elements (mTEs) such as miniature inverted-repeat transposable element (MITE), terminal repeat retrotransposon in miniature, and short interspersed element are exquisite sources for marker development. mTEs are short, non-autonomous and stably inherited. The high-copy members are widely distributed into the gene rich euchromatic regions. Here, we conducted a modified transposon display (TD) for a high-copy MITE family, BraSto-2 (Bs2). The Bs2-specific primers derived from conserved sequences of Bs2 members as well as MseI adapter primers were used for polymerase chain reaction (PCR) in two Brassica rapa accessions, ‘Chiifu’ and ‘Kenshin’. The pooled PCR products were sequenced by Illumina sequencing platform instead of high-resolution gel electrophoresis. Subsequent in silico-based insertion polymorphism (IP) analysis (next-generation sequencing [NGS]-based Bs2 transposon display) was conducted, which generated more than 99 putative polymorphic insertion sites between ‘Chiifu’ and ‘Kenshin’. Among 90 successful PCR amplification, 34 showed Bs2 IP (IP-Bs2) between ‘Chiifu’ and ‘Kenshin’ accessions, 27 and seven ‘Chiifu’- and ‘Kenshin’-unique insertions, respectively. When the 90 IP-Bs2 primer sets were applied to 10 Brassica accessions, including four additional B. rapa and B. oleracea accessions, 69 (76%) showed insertion olymorphism among accessions. The IP-Bs2 were evenly distributed through all the chromosomes and provide rich polymorphism among various B. rapa and B. oleracea accessions demonstrating the usefulness of these markers for various genetic diversity and molecular breeding studies in Brassica. In addition, NGS-based TD will be applicable to various high copy transposable elements family for high throughput and rapid polymorphic marker development which will be helpful for efficient plant genomics and breeding purposes.

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

( Su-jeong Yang ),( Hyun-woo Kim ),( Seok-gwan Choi ),( Sangdeok Chung ),( Seok Jin Oh ),( Shweta Borkar ),( Hak Jun Kim ) 한국미생물 · 생명공학회 2019 Journal of microbiology and biotechnology Vol.29 No.12

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 algaebacteria consortia, which would be useful for various biotechnological applications including wastewater treatment, bioremediation, and sustainable aquaculture.

Toward Complete Bacterial Genome Sequencing Through the Combined Use of Multiple Next-Generation Sequencing Platforms

( Hae Young Jeong ),( Dae Hee Lee ),( Choong Min Ryu ),( Seung Hwan Park ) 한국미생물 · 생명공학회 2016 Journal of microbiology and biotechnology Vol.26 No.1

PacBio’s long-read sequencing technologies can be successfully used for a complete bacterial genome assembly using recently developed non-hybrid assemblers in the absence of secondgeneration, high-quality short reads. However, standardized procedures that take into account multiple pre-existing second-generation sequencing platforms are scarce. In addition to Illumina HiSeq and Ion Torrent PGM-based genome sequencing results derived from previous studies, we generated further sequencing data, including from the PacBio RS II platform, and applied various bioinformatics tools to obtain complete genome assemblies for five bacterial strains. Our approach revealed that the hierarchical genome assembly process (HGAP) non-hybrid assembler resulted in nearly complete assemblies at a moderate coverage of ~75x, but that different versions produced non-compatible results requiring post processing. The other two platforms further improved the PacBio assembly through scaffolding and a final error correction.

Next Generation Sequencing을 통한 미생물 군집 분석의 축산분야 활용

김민석,백열창,오영균,Kim, Minseok,Baek, Youlchang,Oh, Young Kyoon 한국축산환경학회 2015 한국축산시설환경학회지 Vol.21 No.3

The objective of this study was to review application of next-generation sequencing (NGS) to investigate microbiome in the livestock sector. Since the 16S rRNA gene is used as a phylogenetic marker, unculturable members of microbiome in nature or managed environments have been investigated using the NGS technique based on 16S rRNA genes. However, few NGS studies have been conducted to investigate microbiome in the livestock sector. The 16S rRNA gene sequences obtained from NGS are classified to microbial taxa against the 16S rRNA gene reference database such as RDP, Greengenes and Silva databases. The sequences also are clustered into species-level OTUs at 97% sequence similarity. Microbiome similarity among treatment groups is visualized using principal coordinates analysis, while microbiome shared among treatment groups is visualized using a venn diagram. The use of the NGS technique will contribute to elucidating roles of microbiome in the livestock sector.

차세대 염기서열 분석 기법과 활용

권경훈 ( Kyoung Hun Kwon ),정해원 ( Hai Won Chung ) 서울대학교 인구의학연구소 2012 人口醫學硏究論集 Vol.25 No.-

The next generation sequencing (NGS) is faster and a lower cost new sequencing technique getting huge-large scale fragment, called reads, and sequence data than traditional sequencing. NGS technologies include a number of methods that are grouped broadly as template preparation, sequencing and imaging, and data analysis. Recently available next generation sequencer are the GS-FLX (Roche). Genome Analyzer (Illumina), and SOLiD (Life Technologies). However, the NGS produces much shorter sequences and large volumes of reads compared to conventional sequencing, whereby huge amounts of short reads are aligned onto a reference. For fast mapping of the massive amounts of short reads, a lot of mapping algorithms and programs have been developed. In this article, we introduce NGS technique and application of NGS.

Application of Next Generation Sequencing in Laboratory Medicine

Zhong Yiming,Xu Feng,Wu Jinhua,Schubert Jeffrey,Li Marilyn M. 대한진단검사의학회 2021 Annals of Laboratory Medicine Vol.41 No.1

The rapid development of next-generation sequencing (NGS) technology, including advances in sequencing chemistry, sequencing technologies, bioinformatics, and data interpretation, has facilitated its wide clinical application in precision medicine. This review describes current sequencing technologies, including short- and long-read sequencing technologies, and highlights the clinical application of NGS in inherited diseases, oncology, and infectious diseases. We review NGS approaches and clinical diagnosis for constitutional disorders; summarize the application of U.S. Food and Drug Administration-approved NGS panels, cancer biomarkers, minimal residual disease, and liquid biopsy in clinical oncology; and consider epidemiological surveillance, identification of pathogens, and the importance of host microbiome in infectious diseases. Finally, we discuss the challenges and future perspectives of clinical NGS tests.

Next Generation Sequencing and Bioinformatics

Ki-Bong Kim(김기봉) 한국생명과학회 2015 생명과학회지 Vol.25 No.3

매우 빠른 속도로 발전하고 있는 차세대 염기서열 분석 플랫폼과 최신 생물정보학적 분석도구들로 말미암아, 1,000달러 이하의 가격으로 인간 유전체 염기서열을 해독하고자 하는 궁극적인 목표가 조만간 곧 실현될 수 있을것 같다. 차세대 염기서열 분석 분야의 급속한 기술적 진전은 NGS 데이터의 분석과 관리를 위한 통계적 방법과 생물정보학적 분석도구들에 대한 수요를 꾸준히 증대시키고 있다. NGS 플랫폼이 상용화되어 쓰이기 시작한 초창기부터, NGS 데이터를 분석하고 해석하거나, 가시화 해주는 다수의 응용프로그램이나 도구들이 개발되어 활용되어 왔다. 그러나, NGS 데이터의 엄청난 범람으로 데이터 저장, 데이터 분석 및 관리 등에 있어서 해결해야 할 많은 문제들이 부각되고 있다. NGS 데이터 분석은 단편서열과 참조서열간의 서열정렬, 염기식별, 다형성 발견, 쌍단편 서열이나 비쌍단편 서열 등을 이용한 어셈블리 작업, 구조변이 발견, 유전체 브라우징 등을 본질적으로 포함한다. 본 논문은 주요 차세대 염기서열 결정기술과 NGS 데이터 분석을 위한 생물정보학적 분석도구들에 대해 개관적으로 소개하고자 한다. 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.