PacBio scientist Ellen Paxinos discusses a study presented at AGBT that gnerated single-molecule full genome sequencing of HIV 1 from two pairs of linked transmission from a Zambian cohort. Sequencing was done on full-length amplicons from the virus, and clustering accurately placed the virus from each pair together, distinguishing between the two pairs. Paxinos notes that 50 MB of sequence data was generated in less than four hours.
UC Davis’s Bart Weimer describes foodborne pathogens and their proclivity for rapid genome rearrangement. The 100K Pathogen Genome Project he leads is using PacBio long-read sequencing to close genomes and analyze methylation; Weimer reports that his team has already discovered new epigenetic modifications in Salmonella and Listeria with the technology.
How does the PacBio sequencer produce epigenetic data? CSO Jonas Korlach describes how the technology works, which DNA modifications can be detected, and gives examples of kinetic signatures for various modifications and their associated target motifs.
Epigenetics expert Michael Jennings from Griffith University first posited the phasevarion, or the phase variable regulon mechanism in host-adapted pathogens. This mechanism switches expression of multiple genes in a coordinated fashion and has significant implications on pathogen virulence. In his talk, Jennings describes the phasevarion and his use of whole methylome data to rapidly identify methylation targets.
Jonas Korlach, CSO of PacBio, discusses the revival of finished genomes the microbial community will see with long read data, emphasizing that for certain organisms such as rapidly evolving microbes, having a de novo finished genome will be more useful than creating a draft based on a previous related reference genome. Korlach describes two bioinformatic methods from PacBio, a hierarchical genome assembly process (HGAP) and an consensus caller (Quiver), which are used to generate finished genomes from just long-read PacBio data, with final genome sequence accuracies over 99.999%. Korlach demonstrates the ability of PacBio data to generate closed, high-quality de…
In this AGBT plenary talk, Jonas Korlach presented a number of collaborative studies between PacBio and other institutions to make use of highly accurate, long-read sequence data, which has led to a revival of finished genomes. Examples from the infectious disease or pathogen realm included Pertussis, Salmonella, and Listeria, all of which now have closed genomes from PacBio-generated data. Korlach also reported on epigenomic information in Salmonella and Listeria, indicating potential new forms of DNA modifications.
Garth Ehrlich from the Center for Genomic Sciences at Allegheny Singer Research Institute reports on new studies of pneumococcal epigenetics. Streptococcus pneumonia, which causes more than 1.6 million deaths annually, has a highly plastic genome. Methylation analysis with SMRT Sequencing found a novel modification in addition to the expected epigenetic changes.
Peter Evans from the US FDA shares insights on whole-genome sequencing for bacteria of importance to public health. Comparing data across PacBio, 454, and MiSeq sequencers, he says having closed genomes, long reads, and methylation patterns are critical for gleaning comprehensive information about a microbe.
Bart Weimer, a professor at the University of California, Davis, who is leading the 100K Foodborne Pathogen Genome Project, talks about using PacBio sequencing to produce long reads for microbial genomes as well as to study how bacteria use epigenetics to regulate gene expression.
Ulf Gyllensten from Uppsala University used SMRT Sequencing to study multi-drug-resistant bacteria. Time to results was faster than other NGS platforms and generally resulted in complete genome assemblies, even for an organism with a 70% AT-rich genome. He also applied SMRT Sequencing for the characterization of HPV subtypes, important in cervical cancer.
In his AGBT talk, Matthew Blow from the Joint Genome Institute describes high-throughput pipelines to annotate gene function and explore methylation in microbes. He uses transposon sequencing to annotate thousands of genes in bacteria and archaea. Later, he presents a study using SMRT Sequencing to generate complete methylomes for 232 prokaryotes, showing that orphan methylases appear to have a regulatory role.
In this poster presentation, PacBio scientist Ellen Paxinos describes an improved algorithm for circular consensus reads. Using this new algorithm, dubbed CCS2, it is possible to reach arbitrarily high quality across longer insert lengths at a lower cost and higher throughput than Sanger Sequencing. She shows results from the application of CCS2 to the characterization of the HIV-1 K103N drug-resistance associated mutation, which is both important clinically, and represents a challenge due to regional sequence context.
In this AGBT poster, PacBio bioinformatician Matthew Seetin presents a new assembly for Aedes aegypti cell line, the mosquito responsible for spreading viruses like Dengue and Zika. SMRT Sequencing generated a gapless assembly with a contig N50 of 1.4 Mb, compared to 82 kb in the previous assembly. The genome features a number of transposable elements and long tandem repeats.