Distinguish one virus within a crowd
Virus replication and competition produces a complex mixture of variants within each infected host. Resolution of all the variants within a population is critical to understanding evolution, quasispecies dynamics, drug-resistance, and immune escape. Distinguishing variants that may differ by only a handful of SNPs across an entire genome requires both the high accuracy and long read lengths that only SMRT Sequencing can provide.
Confident and complete resolution of viral diversity
Single Molecule, Real-Time (SMRT) Sequencing produces amplicon sequencing reads that average up to 100 kb in length, providing an unprecedented view of complete viral genes and genomes. Highly accurate, single-molecule consensus reads enable direct detection of phased variants, giving you the ability to:
- Deconvolute complex mixtures into quasispecies and unique haplotypes
- Track the evolution and phylogeny of viral populations
- Identify and quantify minor variants
- Generate complete de novo assemblies of large viral genomes
Workflow: from DNA to resolved viral populations
|Sample & Library Preparation|
|Obtain targeted sequences with flexible sample and library workflows.|
|Scale coverage based on your project needs and achieve up 500,000 high-accuracy reads per SMRT Cell 1M on the Sequel System.|
- Maximize output and turn-around-time with adjustable run parameters
- Choose 10 or 20 hour movie time based on amplicon or insert size
*Read lengths, number of reads, data per SMRT Cell, and other sequencing performance results vary based on sample quality/type and insert size, among other factors.
|Make discoveries using bioinformatics tools in SMRT Analysis or PacBio DevNet.|
- Circular Consensus Sequence (CCS): Generate high-fidelity reads by identifying consensus sequences for single DNA molecules
- Long Amplicon Analysis (LAA): generate reference-free, phased consensus assemblies from pooled amplicons
- Minor Variant Analysis: detect and quantify SNPs down to 1% abundance in a heterogenous data set using a reference
- HGAP: create high-quality de novo viral genome assemblies with long sequence continuity and high accuracy (>QV50)
Spotlight: SMRT Sequencing of full-length HIV env reveals complete compartmentalization of variants in brain versus all other tissues
Phylogenetic analysis of high-quality consensus reads shows that brain-derived viruses are compartmentalized and evolutionarily distinct from virus in tissues outside the brain. By generating thousands of full-length HIV env sequences in a single run, the authors definitively show that virus in each of the three brain tissues studied is genetically distinct, whereas variants from all peripheral tissues are dispersed throughpout the tree. Explore this research further:
Brese, R. L., et al. (2018). Ultradeep single-molecule real-time sequencing of HIV envelope reveals complete compartmentalization of highly macrophage-tropic R5 proviral variants in brain and CXCR4-using variants in immune and peripheral tissues. Journal of Neurovirology, ePub ahead of print.
To learn more about how to resolve your viral populations, contact us.
- Kumari, Romika et al. (2018) PausePred and Rfeet: webtools for inferring ribosome pauses and visualizing footprint density from ribosome profiling data. RNA
- Brese, Robin L et al. (2018) Ultradeep single-molecule real-time sequencing of HIV envelope reveals complete compartmentalization of highly macrophage-tropic R5 proviral variants in brain and CXCR4-using variants in immune and peripheral tissues. Journal of neurovirology
- Mehta, Devang et al. (2018) CRISPR-Cas9 interference in cassava linked to the evolution of editing-resistant geminiviruses BioRxiv
- Paulk, Nicole K et al. (2018) Bioengineered AAV capsids with combined high human liver transduction in vivo and unique humoral seroreactivity. Molecular therapy
- Tai, Phillip WL et al. (2018) Adeno-associated virus genome population sequencing achieves full vector genome resolution and reveals human-vector chimeras Molecular therapy
- Imamichi, Hiromi et al. (2016) Defective HIV-1 proviruses produce novel protein-coding RNA species in HIV-infected patients on combination antiretroviral therapy. Proceedings of the National Academy of Sciences of the United States of America
- Laird Smith, Melissa et al. (2016) Rapid sequencing of complete env genes from primary HIV-1 samples Virus evolution
- Bull, Rowena A et al. (2016) A method for near full-length amplification and sequencing for six hepatitis C virus genotypes. BMC genomics
- Dilernia, Dario A et al. (2015) Multiplexed highly-accurate DNA sequencing of closely-related HIV-1 variants using continuous long reads from single molecule, real-time sequencing. Nucleic acids research
- Rogers, Matthew B et al. (2015) Intrahost dynamics of antiviral resistance in influenza a virus reflect complex patterns of segment linkage, reassortment, and natural selection. mBio
- Paulk, Nicole K et al. (2018) Bioengineered viral platform for intramuscular passive vaccine delivery to human skeletal muscle. Molecular therapy
- Ramsay, Joshua D et al. (2018) Hepacivirus A infection in horses defines distinct envelope hypervariable regions and elucidates potential roles of viral strain and adaptive immune status in determining envelope diversity and infection outcome. Journal of virology
- Damilano, Gabriel Dario et al. (2018) Computational comparison of availability in CTL/gag epitopes among patients with acute and chronic HIV-1 infection. Vaccine
- Davidsson, Marcus et al. (2018) Molecular barcoding of viral vectors enables mapping and optimization of mRNA trans-splicing. RNA
- Knyazev, Sergey et al. (2018) CliqueSNV: Scalable reconstruction of intra-host viral populations from NGS reads BioRxiv
- Banga, Riddhima et al. (2018) Blood CXCR3+CD4 T cells are enriched in inducible replication competent HIV in aviremic antiretroviral therapy-treated individuals. Frontiers in immunology
- Yoon, Yeonsoo et al. (2018) Streamlined ex vivo and in vivo genome editing in mouse embryos using recombinant adeno-associated viruses. Nature communications
- Poster: Laird Smith, M. et al. (2016) An improved circular consensus algorithm with an application to detect HIV-1 Drug Resistance Associated Mutations (DRAMs)
- Poster: Hepler, N. Lance et al. (2016) An improved circular consensus algorithm with an application to detection of HIV-1 Drug-Resistance Associated Mutations (DRAMs)
- Poster: Brown, Michael et al. (2015) High-accuracy, single-base resolution of near-full-length HIV genomes.
- Poster: Murrell, Ben et al. (2015) Full-length env deep sequencing in a donor with broadly neutralizing V1/V2 antibodies.
- Poster: Brown, Michael et al. (2015) Sequencing complex mixtures of HIV-1 genomes with single-base resolution.
- Poster: Laird Smith, Melissa et al. (2015) Full-length HIV-1 env deep sequencing in a donor with broadly neutralizing V1/V2 antibodies.
- (2017) Tutorial: Minor variant analysis [SMRT Link v5.0.0]
- Paulin, Lars (2017) AGBT Virtual Poster: Single-molecule sequencing reveals the presence of distinct JC polyomavirus populations in patients with progressive multifocal leukoencephalopathy
- (2017) Tutorial: Circular Consensus Sequence analysis application [SMRT Link v5.0.0]
- PacBio Certified Service Providers (2018)
- SMRT Sequencing Brochure: Revolutionize genomics with SMRT Sequencing (2018)
- SMRT Analysis Brochure: Gain a deeper understanding of your sequencing data (2017)
- Industrial Biotechnology Brochure: Fuel biotech discovery with confident characterization of microbes and their communities. (2016)
- Case Study: Scientists advance understanding of microbial genomes, epigenomes and communities with SMRT Sequencing (2016)
- Certified Service Provider Profile: Institute for Genome Sciences expands long-read sequencing services with new Sequel System (2016)