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Tuesday, June 1, 2021

A method for the identification of variants in Alzheimer’s disease candidate genes and transcripts using hybridization capture combined with long-read sequencing

Alzheimer’s disease (AD) is a devastating neurodegenerative disease that is genetically complex. Although great progress has been made in identifying fully penetrant mutations in genes such as APP, PSEN1 and PSEN2 that cause early-onset AD, these still represent a very small percentage of AD cases. Large-scale, genome-wide association studies (GWAS) have identified at least 20 additional genetic risk loci for the more common form of late-onset AD. However, the identified SNPs are typically not the actual risk variants, but are in linkage disequilibrium with the presumed causative variant (Van Cauwenberghe C, et al., The genetic landscape of Alzheimer disease: clinical…

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Tuesday, June 1, 2021

A method for the identification of variants in Alzheimer’s disease candidate genes and transcripts using hybridization capture combined with long-read sequencing

Alzheimer’s disease (AD) is a devastating neurodegenerative disease that is genetically complex. Although great progress has been made in identifying fully penetrant mutations in genes such as APP, PSEN1 and PSEN2 that cause early-onset AD, these still represent a very small percentage of AD cases. Large-scale, genome-wide association studies (GWAS) have identified at least 20 additional genetic risk loci for the more common form of late-onset AD. However, the identified SNPs are typically not the actual causal variants, but are in linkage disequilibrium with the presumed causative variant (Van Cauwenberghe C, et al., The genetic landscape of Alzheimer disease: clinical…

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Tuesday, June 1, 2021

Characterizing the pan-genome of maize with PacBio SMRT Sequencing

Maize is an amazingly diverse crop. A study in 20051 demonstrated that half of the genome sequence and one-third of the gene content between two inbred lines of maize were not shared. This diversity, which is more than two orders of magnitude larger than the diversity found between humans and chimpanzees, highlights the inability of a single reference genome to represent the full pan-genome of maize and all its variants. Here we present and review several efforts to characterize the complete diversity within maize using the highly accurate long reads of PacBio Single Molecule, Real-Time (SMRT) Sequencing. These methods provide…

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Tuesday, June 1, 2021

Microbiome profiling at the strain level using rRNA amplicons

Strain level microbiome profiling is needed for a full understanding of how microbial communities influence human health. Microbiome profiling of rRNA gene amplicons is a well-understood method that is rapid and inexpensive, but standard 16S rRNA gene methods generally cannot differentiate closely related strains. Whole genome/shotgun microbiome profiling is considered a higher-resolution alternative, but with decreased throughput and significantly increased sequencing costs and analysis burden. With both methods there are also challenges with microbial lysis, DNA preparation, and taxonomic analysis. Specialized microbiome-focused protocols were developed to achieve strain-level taxonomic differentiation using a rapid, high throughput rRNA gene assay. The protocol…

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Friday, February 5, 2021

Webinar: A paradigm shift in HLA sequencing: from exons to high-resolution allele-level HLA yyping

Human MHC class I genes HLA-A, -B, -C, and class II genes HLA -DR, -DQ, and -DP play a critical role in the immune system as primary factors responsible for organ transplant rejection. Additionally, the HLA genes are important targets for clinical and drug sensitivity research because of their direct or linkage-based association with several diseases, including cancer, and autoimmune diseases. HLA genes are highly polymorphic, and their diversity originates from exonic combinations as well as recombination events. With full-length gene sequencing, a significant increase of new alleles in the HLA database is expected, stressing the need for high-resolution sequencing.…

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Friday, February 5, 2021

Webinar: Amplicon sequencing with confidence – High-fidelity, long-read PacBio sequencing solutions

In this webinar, Lori Aro and Cheryl Heiner of PacBio describe how high-throughput amplicon sequencing using Single Molecule, Real-Time (SMRT) Sequencing and the Sequel System allows for the easy and cost-effective generation of high-fidelity, long reads from amplicons ranging in size from several hundred base pairs to 20 kb. Topics covered include the latest advances in SMRT Sequencing performance for detection of all variant types even in difficult to sequence regions of the genome, multiplexing options to increase throughput and improve efficiency, and examples of amplicon sequencing of clinically relevant targets.

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Friday, February 5, 2021

PAG Conference: How SMRT Sequencing is accelerating plant and animal genomics

In this presentation, Justin Blethrow provides an overview of recent and upcoming developments across PacBio’s SMRT Sequencing product portfolio, and their implications for PacBio’s major applications. In presenting the product roadmap, he illustrates how key new products coming in 2019 will make SMRT Sequencing dramatically more affordable and easy to use, and how they will enable customers to routinely produce highly accurate, single-molecule long reads.

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Friday, February 5, 2021

PAG Conference: The impact of highly accurate PacBio sequence data on the assembly of a tetraploid rose

In this presentation at PAG 2020, Bart Nijland of Genetwister Technologies explains how his team set out to make a haplotype-aware assembly of the highly complex tetraploid Rosa x hybrida L. genome in order to capture its full range of genetic variation. HiFi reads generated from PacBio’s Sequel II System have made it possible to parse out critical information from many of the plant’s parental genes.

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Friday, February 5, 2021

AGBT Presentation: Feed the World – Developing genomic resources for insects as food

In a push to develop insect-based food sources for people, Brenda Oppert from the USDA has been sequencing bug genomes with PacBio technology. Long reads are essential because of the highly repetitive sequences and large genomes. On the Sequel II System, a single SMRT Cell is sufficient to generate 350-fold coverage and produce a high-quality assembly for some of the insects she’s studying.

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Tuesday, April 21, 2020

The bracteatus pineapple genome and domestication of clonally propagated crops.

Domestication of clonally propagated crops such as pineapple from South America was hypothesized to be a ‘one-step operation’. We sequenced the genome of Ananas comosus var. bracteatus CB5 and assembled 513?Mb into 25 chromosomes with 29,412 genes. Comparison of the genomes of CB5, F153 and MD2 elucidated the genomic basis of fiber production, color formation, sugar accumulation and fruit maturation. We also resequenced 89 Ananas genomes. Cultivars ‘Smooth Cayenne’ and ‘Queen’ exhibited ancient and recent admixture, while ‘Singapore Spanish’ supported a one-step operation of domestication. We identified 25 selective sweeps, including a strong sweep containing a pair of tandemly duplicated…

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Tuesday, April 21, 2020

Characterization of Reference Materials for Genetic Testing of CYP2D6 Alleles: A GeT-RM Collaborative Project.

Pharmacogenetic testing increasingly is available from clinical and research laboratories. However, only a limited number of quality control and other reference materials currently are available for the complex rearrangements and rare variants that occur in the CYP2D6 gene. To address this need, the Division of Laboratory Systems, CDC-based Genetic Testing Reference Material Coordination Program, in collaboration with members of the pharmacogenetic testing and research communities and the Coriell Cell Repositories (Camden, NJ), has characterized 179 DNA samples derived from Coriell cell lines. Testing included the recharacterization of 137 genomic DNAs that were genotyped in previous Genetic Testing Reference Material Coordination…

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Tuesday, April 21, 2020

Identification and characterization of chicken circovirus from commercial broiler chickens in China.

Circoviruses are found in many species, including mammals, birds, lower vertebrates and invertebrates. To date, there are no reports of circovirus-induced diseases in chickens. In this study, we identified a new strain of chicken circovirus (CCV) by PacBio third-generation sequencing samples from chickens with acute gastroenteritis in a Shandong commercial broiler farm in China. The complete genome of CCV was verified by inverse PCR. Genomic analysis revealed that CCV codes two inverse open reading frames (ORFs), and a potential stem-loop structure was present at the 5′ end with a structure typical of a circular virus. Phylogenetic tree analysis showed that…

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Tuesday, April 21, 2020

Chromosome-length haplotigs for yak and cattle from trio binning assembly of an F1 hybrid

Background Assemblies of diploid genomes are generally unphased, pseudo-haploid representations that do not correctly reconstruct the two parental haplotypes present in the individual sequenced. Instead, the assembly alternates between parental haplotypes and may contain duplications in regions where the parental haplotypes are sufficiently different. Trio binning is an approach to genome assembly that uses short reads from both parents to classify long reads from the offspring according to maternal or paternal haplotype origin, and is thus helped rather than impeded by heterozygosity. Using this approach, it is possible to derive two assemblies from an individual, accurately representing both parental contributions…

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Tuesday, April 21, 2020

An improved pig reference genome sequence to enable pig genetics and genomics research

The domestic pig (Sus scrofa) is important both as a food source and as a biomedical model with high anatomical and immunological similarity to humans. The draft reference genome (Sscrofa10.2) represented a purebred female pig from a commercial pork production breed (Duroc), and was established using older clone-based sequencing methods. The Sscrofa10.2 assembly was incomplete and unresolved redundancies, short range order and orientation errors and associated misassembled genes limited its utility. We present two highly contiguous chromosome-level genome assemblies created with more recent long read technologies and a whole genome shotgun strategy, one for the same Duroc female (Sscrofa11.1) and…

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Tuesday, April 21, 2020

A robust benchmark for germline structural variant detection

New technologies and analysis methods are enabling genomic structural variants (SVs) to be detected with ever-increasing accuracy, resolution, and comprehensiveness. Translating these methods to routine research and clinical practice requires robust benchmark sets. We developed the first benchmark set for identification of both false negative and false positive germline SVs, which complements recent efforts emphasizing increasingly comprehensive characterization of SVs. To create this benchmark for a broadly consented son in a Personal Genome Project trio with broadly available cells and DNA, the Genome in a Bottle (GIAB) Consortium integrated 19 sequence-resolved variant calling methods, both alignment- and de novo assembly-based,…

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