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Next Generation Sequencing

GTCbio | September 8th, 2017

Nucleotide sequencing is the method used for determining the precise order of nucleotides within a DNA/RNA molecule. The identification of complete gene and the complete genome of Bacteriophage MS2 in 1972 and 1976 respectively; and the invention of the first method for DNA sequencing involving a location-specific primer extension strategy in 1970 were the major landmarks of RNA and DNA sequencing. The pioneer of sequencing was Frederick Sanger, who was awarded two Nobel prizes, the first for the sequencing of proteins, and the second for the sequencing of DNA. The field of sequencing saw a lot of development in the mid to late 1990s with the emergence of high-throughput sequencing (HTP) methods including “next-generation” and “second-generation” sequencing methods. Next-generation sequencing (NGS), with its unprecedented throughput, scalability, and speed, has become a staple for researchers in biological systems studies. Using NGS thousands or millions of sequences can be concurrently produced in a single sequencing process. NGS also allows rapid sequencing of a whole genome, exome sequencing, and de novo sequencing. It can also be used for RNA sequencing to discover novel RNA variants, splice sites, or for precisely quantifying mRNAs for gene expression analysis. In addition, NGS has application for analysis of epigenetic factors including genome-wide DNA methylation and DNA-protein interactions, the sequencing of cancer samples to study rare somatic variants, and tumor subclones. Microbial diversity in humans, targeted sequencing, ChIP (chromatin immunoprecipitation) sequencing; and ribosome profiling are all being facilitated by NGS.

Methods using a pool of DNA or RNA from thousands to millions of cells result in findings that may not be reflective of individual cells from within that population. The advances in have overcome this drawback with resulting in accurate determination of information about the genomic and transcriptomic profiles of single cells. In the last decade there have been significant advances in the capability and availability of high resolution and high throughput sequencing of single cell genomes. Single-cell analysis reduces the complexity of the genomic signal through the physical separation of cells or chromosomes providing higher-resolution views of the genomic content of samples. Recent research suggests that the accuracy of transcriptional profiles produced from single cells does not seem to suffer significantly if samples used are frozen. RNA and DNA sequencing involving single cell analysis has opened many doors including the study of un-culturable microorganisms; populations of rare cells; and individual cells within bulk populations. Some of the techniques include RNA-seq, whole genome sequencing, whole-exome sequencing, whole-transcriptomic sequencing, epigenomic sequencing. Various technologies exploiting single cell sequencing include Fluidigm technologies, inDrop; DropSeq, ICell8 and CellRaft. Single cell sequencing is being used in cancer studies, immune system studies, prenatal diagnosis and neurobiological studies.

It is forecasted that next generation sequencing clinical diagnostic market will see continued rapid growth over the next several years. The power of exome sequencing for clinical diagnostics has been exploited in recent time since the exome is to the location of 85% of disease-causing genetic variants. It has helped in highly accurate diagnosis of rare hereditary disorders. Sophia Genetics has released Whole Exome Solution (WES) and Clinical Exome Solution (CES) that are accessible through Sophia DDM® (analytical platform for clinical diagnostic). NGS increasingly used in the field of personalized medicine and companion diagnostics. One of the latest diagnostic sequencing kits includes Illumina’s FDA-approved next-generation sequencing cancer companion diagnostic test kit. Recent research suggests that transcriptome sequencing may provide diagnoses for patients with Mendelian disorders. An upcoming field may be use of whole-genome sequencing in conjunction with artificial intelligence-based analysis to quickly identify clinically actionable mutations associated with brain tumors. Other applications include cancer diagnostics, including NGS-based tests for oncology, liquid biopsies, and detection of clinically relevant mutations using 5 circulating tumor cells (CTCs). NGS will also find use in the genetic analysis ofinherited disorders, prenatal and postnatal diagnostics; infectious disease diagnostics; human leukocyte antigen (HLA) testing; and in the direct-to-consumer genetic testing industry.

Next Generation Sequencing

The GTCbio 7thGenomics and Big Data Summit Conference will be held on September 26-27, 2017 in San Diego, CA and the speakers for the session entitled “Next Generation Sequencing” include the following:

     •  John Oliver, Chief Technology Officer, Nabsys 2.0

Dr. John Oliver’s is responsible for overseeing all aspects of the scientific development w.r.t. DNA mapping platform. His diverse experience includes synthetic organic chemistry, biochemistry, molecular biology, surface chemistry, and microarray technology.

     •  Flora Tassone, Professor, Biochemistry and Molecular Medicine; M.I.N.D. Institute, Investigator, University of California, Davis, School of Medicine

Dr. Flora’s expertise includes transcriptional and translational regulation of the fragile X (FMR1) gene. Dr. Flora has been involved in the development of molecular biomarkers predictive of drug efficacy and monitoring disease severity. Her extensive experience includes medical genetics and clinical analysis.

     •  Joshua Edel, Professor of Biosensing & Analytical Sciences, Imperial College London

Dr. Joshua’s research areas include development of single molecule detection within microfluidic systems; nanobiotechnology; single molecule biophysics. In 2016, Dr. Joshua was awarded prestigious ERC Starting Grant on “Nanoporous Membranes for High Throughput Rare Event Bioanalysis”.

     •  Michael Phelan, Senior Scientist, R&D, Fluidigm Corporation

Dr. Michael Phelan is actively working to develop NGS library prep on Fluidigm instrument platforms.

     •  Richard McCombie, Professor & Director, The Stanley Institute for Cognitive Genomics, Cold Spring Harbor Laboratory

Dr. Richard harnesses revolutionary improvements in DNA sequencing technology to assemble genomes for a variety of organisms and probe the genetic basis of neurological disorders, including autism and schizophrenia; and better understand cancer progression and understand the complex structures of the genomes of higher plants.

     •  Tyson Clark, Director, Applications Development, Pacific Biosciences

Dr. Clark focuses on development of new applications that utilize PacBio’s long read sequencing technologies. He was a pioneer in the use of microarrays to study alternative splicing at a genome-wide scale. He has worked on a broad range of applications of SMRT® Sequencing, including direct detection of modified DNA bases.

     •  Meni Wanunu, Professor, Department of Physics, Northeastern University

Dr. Meni is involved in studies related to biosystems at the nanoscale (macromolecular and sub-molecular levels). His group is developing novel techniques that probe how small molecular changes affect the global properties of macromolecules and biomolecules. Using various tools enabled by nanotechnology, the group investigates biomolecular structure and dynamics at their corresponding size scale.

     •  John Thompson, Chief Technology Officer, Claritas Genomics

Dr. John’s career has spanned around pharmaceutical, next-gen sequencing, and genetic testing industries. He is responsible for improving genetic assays for diagnosing pediatric disease and developing the next generation of assays. He has used molecular biology and genetic approaches to identify novel therapeutic targets, to establish the genetic basis of drug response, and to characterize the genetics of drug-induced adverse events.

We invite you to join us at the 7th Genomics and Big Data Summit in San Diego on September 26-27 at the Coronado Island Marriott Resort.