DNA sequencing

Summary

This report briefly reviews basics of human genome variations, development of sequencing technologies, and their applications. Current large and small sequencers are described as well as companies developing them. Various applications of sequencing are described including those for genetics, medical diagnostics, drug discovery and cancer. Next generation sequencing technologies, both second and third generations, are reviewed. Companies developing software for analysis of sequencing data are also included. Selected academic institutes conducting research in sequencing are also listed.

Current market is mostly for research applications and future markets will be other applications related to healthcare. The value of DNA sequencer market in 2009 is described with estimates for 2014 and 2019. Various methods and factors on which market estimates depend are described briefly. Small sequencers form the basis of SWOT (strengths, weaknesses, opportunities, threats) analysis. Several marketing strategies have been outlined.

The report includes profiles of 54 companies involved in sequencing and their 47 collaborations. The report text is supplementd by 28 tables, 8 figures and 160 selected references to the literature.

Contents

• 0. Executive Summary
• 1. Introduction
  • Definition and scope of sequencing
  • Historical aspects of sequencing
  • Basics of molecular biology
  • DNA
  • DNA polymerases
  • Restriction endonucleases
  • DNA methylation
  • RNA
  • RNA polymerases
  • Non-coding RNAs
  • DNA transcription
  • Chromosomes
  • Telomeres
  • Mitochondrial DNA
  • Genes
  • The genetic code
  • Gene expression
  • The human genome
  • Variations in the human genome
  • Variations in DNA sequences
  • Single nucleotide polymorphisms
  • Complex chromosomal rearrangements
  • Insertions and deletions in the human genome
  • Large scale variation in human genome
  • Variation in copy number in the human genome
  • Structural variations in the human genome
  • Mapping and sequencing of structural variation from human genomes
  • Impact of sequencing on healthcare
• 2. DNA Sequencing Technologies
  • Introduction
  • DNA extraction and sample preparation
  • Electrophoresis-based method
  • Microfluidics-based extraction and samnple preparation
  • Pressure Cycling Technology
  • Selective immobilization of nucleic acids onto magnetic microparticles
  • Targeted and hybridization-based DNA capture
  • Sanger-sequencing technology
  • Dye-terminator sequencing
  • Large-scale sequencing
  • Automated DNA-sequencing
  • Enhancements of Sanger-sequencing
  • ABI PRISM® 310 Genetic Analyzer
  • Limitations of current sequencing methods and measures to remedy them
  • High-throughput paired end transcriptome sequencing
  • Emerging sequencing technologies
  • Chemical DNA sequencing
  • Second generation sequencers
  • 4300 DNA analyzer
  • Apollo 100
  • Applied Biosystems 3500 series Genetic Analyzer
  • "Color blind" approach to DNA sequencing
  • Cyclic array sequencing
  • CEQ™ 8000
  • DeepCAGE sequencing
  • Electron microscope-based DNA sequencing
  • GenometricaÔ sequencer
  • GS-FLEX system (Roche/454)
  • IBS sequencing technology
  • Illumina Genome Analyzer System
  • Ion Torrent's sequencing technology
  • MegaBACE 500
  • Microdroplet-based PCR for large-scale targeted sequencing
  • Multiplex amplification of human DNA sequences
  • Nanoscale sequencing
  • Polonator sequencer
  • RainStorm™ microdroplet technology
  • Sequential DEXAS
  • SOLiD system: sequencing by ligation
  • PCR-based DNA sequencing technologies
  • Digital PCR
  • Emulsion PCR
  • Bridge amplification PCR system
  • Dual primer emulsion PCR
  • Nucleic acid sequence-based amplification
  • Microarray-based DNA sequencing technologies
  • Arrayit's® H25K
  • High-throughput array-based resequencing
  • Human exome microarrays
  • Optical Mapping
  • Sequencing by hybridization
  • SOLiD-System based ChIP-Sequencing
  • Companies developing whole genome chips/microarrays
  • Next generation sequencing vs microarrays for gene expression profiling
  • Third generation sequencing
  • SOLiD4 System
  • SOLiD PI System
  • Detection of single molecules for sequencing
  • DNA sequence by use of nanoparticles
  • Helicos™ Genetic Analysis System
  • Molecular Combing
  • Nanopore sequencing
  • PNA-based single-molecule detection of specific DNA sequences
  • Sequencing-by-synthesis for single-molecule sequencing
  • Single molecule DNA sequencing by use of carbon nanotubes
  • Single molecule sequencing using Qdot nanocrystals
  • Single-molecule DNA sequencing in a sTOP chip nanowell
  • Single-molecule real-time sequencing
  • Single cell DNA sequencing
  • Role of bioinformatics in sequencing
  • Growth of the sequencing database
  • Data storage
  • Bioinformatics challenges of new sequencing technology
  • Bioinformatic tools for analysis of genomic sequencing data
  • Commercially available software for DNA sequencing
  • ChIA-PET tool for analysis with paired-end tag sequencing
  • Detection of CNVs and gene duplications
  • Sequencing data storage
  • Sequencing analysis tools at academic organizations
  • Accessing DNA sequence information
• 3. Comparative Analysis of Sequencing Technologies
  • General findings of the study
  • Sanger versus second generation marketed sequencers
  • Common features and differences among second generation sequencers
  • Third generation large sequencers
  • SOLiD4 versus competing large sequencers
  • Illumina's HiSeq 2000
  • The ideal small sequencer
  • SWOT analysis of small sequencers
  • Concluding remarks on SWOT analysis
• 4. Research Applications of sequencing
  • Introduction
  • Applications in basic research
  • ChIA-PET technology for 3D study of the genome
  • ChIP-Seq for study of gene expression in embryonic stem cells
  • Chromatin profiling by direct DNA sequencing
  • Epigenetic modifications analyzed by next generation sequencing
  • Genome sequencing with combinatorial probe anchor ligation
  • GS-FLX sequencing for simultaneous detection of mutation and CNV
  • Identifying protein-coding genes in genomic sequences
  • mRNA sequencing
  • Mutation rate measured by direct sequencing
  • Sequencing the transcriptomes of stem cells
  • Synthetic sequence in a bacterial cell
  • Sequencing of human genomes
  • Whole genome sequencing
  • Whole genome resequencing
  • Personal genome sequencing
  • Missing human genome sequences
  • Role of sequencing in identification of human remains
  • Sequence map of the human pan-genome
  • Sequencing of African genomes
  • Sequencing mitochondrial genome
  • Sequencing of ancient genomes
  • Sequencing genomes of other organisms
  • Sequencing of the mouse genome
  • Sequencing of the dog genome
  • Sequencing of the frog genome
  • Sequencing genomes of non-human primates
  • Applications in drug discovery and development
  • Resequencing
  • RNA profiling
  • Transcriptome sequencing for mRNA Expression
  • RNA splice variants
  • NHGRI's sequencing initiatives
  • Approved medical sequencing projects
  • 1000 Genomes Project
  • HapMap catalog as a foundation
  • Role of SOLiD™ System in 1000 Genomes Project
  • Concluding remarks
  • Human Variome Project
  • Academic centers conducting research on sequencing
• 5. Applications of Sequencing in Healthcare
  • Introduction
  • Applications of sequencing in molecular diagnostics
  • Cancer
  • Assessment of sequencing technologies for analyzing tumor DNA
  • Cancer Genome Atlas
  • Detection of cancer biomarkers
  • Discovery of biomarkers for personalizing cancer treatment
  • Discovery of amplified cancer genes
  • Digital proteomics for cancer profiling
  • Epigenome profiling
  • Insights into mutational processes
  • Paired-end sequencing
  • RNA-Seq to study cancer transcriptome
  • Sequencing cancer cell lines
  • Sequencing in acute myeloid leukemia
  • Sequencing brain tumors
  • Sequencing in breast cancer
  • Sequencing of complex human cancer genomes
  • Sequencing mitochondrial DNA to identify cancer biomarkers
  • Sequencing to guide cancer management
  • Genetic disorders
  • Array-based whole-exome sequencing in Bartter syndrome
  • DNA sequencing of maternal plasma
  • Exome sequencing for discovery of the gene for Miller syndrome
  • Whole genome sequencing in Charcot-Marie-Tooth disease
  • Sequencing in Huntington's disease
  • Sequencing genomes of the newborn to screen for genetic disorders
  • Study of rare variants in pinpointing disease-causing genes
  • Sequencing genomes of microbes
  • DNA sequencing for study of bacterial epidemics
  • Next generation sequencing for antibacterial drug discovery
  • Sequencing for mapping genomic variation in Mycobacterium ulcerans
  • Sequencing for mapping genetic interactions in bacteria
  • Sequencing in the management of HIV/AIDS
  • Sequencing genome of Lassa fever virus
  • Future prospects of next generation sequencing
  • Devices for next generation sequencing
  • COLD-PCR and sequencing
  • Next generation sequencing for population targeted sequencing studies
  • Next generation sequencing and drug design
  • Next-generation sequencing in aging research
  • Applications of next generation sequencing in molecular diagnostics
  • Next generation sequencing and personalized medicine
• 6. Markets for Sequencers
  • Introduction
  • Methods used for estimation of sequencer markets
  • Currently marketed sequencers
  • Academic and research markets for sequencing
  • Factors affecting future development of sequencing markets
  • Future needs and support of research
  • Bioinformatics in relation to sequencing
  • Reducing the cost of human genome sequencing
  • US Government-supported research on sequencing
  • Genome X Prize Foundation
  • Innovations to reduce cost of whole genome sequencing
  • Commercial prospects of low cost genome sequencing
  • Genome sequencing suitable for personalized medicine
  • The global sequencing market
  • Marketing potential for sequencers
  • Challenges to developing market for sequencers
  • Recommendations
• 7. Companies Involved in Sequencing
  • Introduction
  • Collaborations
• 8. References
• Tables
  • Table 1-1: Historical landmarks in DNA sequencing
  • Table 1-2: Genetic variations in the human genome
  • Table 2-1: ChIP detection platforms for sequencing
  • Table 2-2: Companies developing whole genome chips/microarrays
  • Table 2-3: Systems for single molecule sequencing
  • Table 2-4: Companies providing DNA sequencing software
  • Table 3-1: Comparison of a generation I and generation II sequencers
  • Table 3-2: Similarities and differences between second generation sequencers
  • Table 3-3: SWOT of ABI 310
  • Table 3-4: SWOT of IBS sequencing
  • Table 3-5: SWOT of NABsys' Hybridization-Assisted Nanopore Sequencing
  • Table 3-6: SWOT of 4300 DNA Analysis System Li-Cor
  • Table 3-7: SWOT of Genometrica
  • Table 3-8: SWOT of Polonator
  • Table 3-9: SWOT of GS FLEX Junior
  • Table 3-10: SWOT of Oxford Nanopore Technologies' nanopore DNA sequencing
  • Table 3-11: SWOT of Ion Torrent™ non-optical sequencer
  • Table 3-12: SWOT of Pacific BioSciences' single-molecule real-time sequencing
  • Table 4-1: Approved medical sequencing projects
  • Table 4-2: Academic centers conducting research on DNA sequencing
  • Table 6-1: Marketed next generation sequencers
  • Table 6-2: De novo sequencing vs resequencing markets
  • Table 6-3: Global markets for sequencing services according to geographical regions
  • Table 6-4: Global markets for sequencing services according to applications
  • Table 6-5: Global markets for sequencers
  • Table 7-1: Companies involved in sequencing
  • Table 7-2: Selected collaborations for DNA sequencing
• Figures
  • Figure 2-1: DNA sequencing process
  • Figure 2-2: Comparison of traditional sequencing and next generation sequencing
  • Figure 2-3: Watson-Crick base pairing
  • Figure 2-4: Workflow of Genome Sequenser FLX system
  • Figure 2-5: Sequencing by ligation
  • Figure 2-6: Construction of SOLiD fragment library using DNA enrichment by ChIP
  • Figure 2-7: Single molecule, realtime DNA sequencing
  • Figure 6-1: Global markets for sequencing services according to applications

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