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The Journey through the Gene: a Focus on Plant Anti-pathogenic Agents Mining in the Omics Era
Published in Mahendra Rai, Chistiane M. Feitosa, Eco-Friendly Biobased Products Used in Microbial Diseases, 2022
José Ribamar Costa Ferreira-Neto, Éderson Akio Kido, Flávia Figueira Aburjaile, Manassés Daniel da Silva, Marislane Carvalho Paz de Souza, Ana Maria Benko-Iseppon
The SGS term describes platforms generating a huge amount (even billions of reads) of short nucleotide sequences (25 to 400 bp), increasing the high-throughput production, and reducing the cost/million bases by orders of magnitude (Mardis 2008). The pyrosequencing system developed by 454 Life Sciences, after acquired by Roche®, was the first successful NGS/SGS platforms. Other successful SGS platforms comprise the Illumina® Genome Analyzer (GA) II/IIx, the Applied Biosystems SOLiD™ (Sequencing by Oligo Ligation Detection), the Helicos HeliScope™, among others (Mardis 2008). Further optimization has led to innovative third-generation (long-read sequencing) platforms as single-molecule real-time sequencing by PacBio®, nanopore sequencing, etc. (Bleidorn 2015; Xiao and Zhou 2020). The sequencing of entire genomes gives rise to a new genetics sub area, the genomics.
“Omics” Technologies in Vaccine Research
Published in Mesut Karahan, Synthetic Peptide Vaccine Models, 2021
The genomic era is a revolution in vaccine development. It started with the shotgun sequencing technology producing the genome sequence of Haemophilus influenzae in 1995 by The Institute for Genomic Research (TIGR). Later, many technologies were developed for next-generation sequencing, such as massively parallel signature sequencing, polony sequencing, 454 pyrosequencing, reversible terminator sequencing, sequencing by oligonucleotide ligation detection (SOLiD), single-molecule real-time sequencing, ion torrent sequencing, or DNA nanoball sequencing (reviewed by Rajesh and Jaya 2017). Due to these advanced technologies, the number of published bacterial genomes increased considerably with 14,754 completed and 128,146 permanent draft genomes deposited in the Genomes Online Database (GOLD, https://gold.jgi.doe.gov) by June 2020. The organisms with completed genomes cover various bacterial pathogens, and they serve vaccine development to find out potential antigens (Serruto et al. 2009).
The Immunoglobulin Variable-Region Gene Repertoire and Its Analysis
Published in Cliburn Chan, Michael G. Hudgens, Shein-Chung Chow, Quantitative Methods for HIV/AIDS Research, 2017
Thomas B. Kepler, Kaitlin Sawatzki
NGS involves the binding of sample DNA molecules to a solid surface followed by cluster amplification to increase the independent molecule readout signal (fluorescence, light, pH). Massive parallel sequencing is conducted by covering the surface in a specific nucleotide and making a call for that base if there is a resulting chemical or physical signal indicating an incorporation event. The nucleotide solution is washed off and the process is repeated, resulting in a large number of sequencing reads for analysis. This process is termed sequencing by synthesis. The most dominant platforms for NGS and Ig-seq are currently the Illumina-based HiSeq and MiSeq sequencers. Similar platforms include the Roche 454 GS series and Ion Torrent PGM. Other NGS approaches include nanopore-based (MinION, Oxford Nanopore Technologies), sequencing by ligation (SOLiD, Life Technologies) and single molecule real-time sequencing (PacBio RS, Pacific Biosciences).
Exploiting differential RNA splicing patterns: a potential new group of therapeutic targets in cancer
Published in Expert Opinion on Therapeutic Targets, 2018
Nidhi Jyotsana, Michael Heuser
The human transcriptome is much more complex than once anticipated, but our growing capacity to define it with more precision and depth using deep sequencing and improved experimental methods have broadened our understanding of splicing in cancer pathogenesis, and helped to define a therapeutic window for targeting. However, most of the currently used sequencing technologies sequence only short stretches of RNA. Therefore, novel technologies are being established such as single molecule real-time sequencing (SMRT by PacBio) [161] or nanopore sequencing [162], which allow sequencing of complete RNA strands and will therefore open up a new dimension of splicing complexity.
Detecting rare thalassemia in children with anemia using third-generation sequencing
Published in Hematology, 2023
Zhen-min Ren, Wu-jiao Li, Zhi-hao Xing, Xiao-ying Fu, Ju-yan Zhang, Yun-sheng Chen, De-fa Li
Single-molecule real-time sequencing (SMRT sequencing) is a third-generation sequencing technology that utilizes a specialized sequencing platform and chemical methods. This method relies on fluorescence signals emitted by DNA polymerase during the amplification process for sequencing.
Molecular techniques for the genomic viral RNA detection of West Nile, Dengue, Zika and Chikungunya arboviruses: a narrative review
Published in Expert Review of Molecular Diagnostics, 2021
Antonio Mori, Elena Pomari, Michela Deiana, Francesca Perandin, Sara Caldrer, Fabio Formenti, Manuela Mistretta, Pierantonio Orza, Andrea Ragusa, Chiara Piubelli
Over the years, the high-throughput NGS platforms showed an epoch-making revolution producing the second generation (SGS), and more recently, the third-generation sequencing (TGS) [149]. NGS is a very helpful tool on the virus discovery and identification in a heterogeneous mix of genetic materials, since it carries out high sensitive massively parallel sequencing and makes doable to analyze data in a sequence-independent manner. According the sequencing technology, the SGS platforms can be regrouped in sequencing by hybridization and sequencing by synthesis (SBS). Currently, SBS approach is the most used by Illumina, the major player in the SBS arena. The main TGS manufacturers are the Pacific Biosciences and the Oxford Nanopore. The Pacific Biosciences platforms (RS and RSII) rely on single molecule real-time sequencing approach (SMRT). The Oxford Nanopore sequencers (the portable MinION, the benchtop GridION, and high throughput PromethION) instead use a protein nanopore-based sequencing. The major bottle neck for NGS is the need of bioinformatic tools with extensive storage and high computational power to analyze and interpretate NGS data. At moment, among the different NGS approaches and platforms, the MinION sequencer is the most cited, especially in field clinical application. MinION is a promising NGS platform suitable to use in remote areas as it is a portable device be able to read multiplexed samples, and cheap having lower initial costs than other sequencers. This platform can also be combined with isothermal assays, such as RT-LAMP. For example, Hayashida et al. [150] developed a RT-LAMP combined with MinION system applicable for both diagnosis of CHIKV infected patients and CHIKV genotyping. The applied IA assay was a dried single-tube RT-LAMP assay (named CZC-LAMP, previously used for the diagnosis of other pathogens [151]), tailored for the specific detection of CHIKV (CHIKV-CZC-LAMP) directly in whole blood or plasma, and the LAMP products then sequenced by MinION. In addition to CHIKV-SL10571 and -S27 laboratory strains, the system was also assessed on clinical samples from recent CHIKV outbreaks in Brazil. Overall, CHIKV-CZC-LAMP showed an analytical sensitivity of <50 PFU/reaction. The consensus generated by MinION sequencing showed a perfect match (100%) with CHIKV-SL10571 and CHIKV-S27 sequences deposited in GenBank, and the phylogenetic analysis that all sequences from CHIKV patients clustered within ECSA genotype, confirming the circulation of the ECSA genotype in Rio de Janeiro during the epidemics in 2016 and 2018. The combination of LAMP assay with MinION was also used by Yamagishi et al. [152] to detect and serotype DENV directly in the field. This combined assay was tested on a total of 233 clinical samples from countries in Southeast Asia with good performance results.