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Application Note: Microbial multiplexing workflow on the Sequel System

2018

Summary +

Obtaining microbial genomes with the highest accuracy and contiguity is extremely important when exploring the functional impact of genetic and epigenetic variants on a genome-wide scale. A comprehensive view of the bacterial genome, including genes, regulatory regions, IS elements, phage integration sites, and base modifications is vital to understanding key traits such as antibiotic resistance, virulence, and metabolism. SMRT Sequencing provides complete genomes, often assembled into a single contig. Our streamlined microbial multiplexing procedure for the Sequel System, from library preparation to genome assembly, can be completed with less than 12 hours bench time. Starting with high-quality genomic DNA (gDNA), samples are sheared to a 10 kb distribution, ligated with barcoded adapters, pooled at equimolar representation, and sequenced. Demultiplexing of samples is automated, allowing for immediate genome assembly on our SMRT Link analysis software solution. The workflow supports up to 16-plex of de novo microbial genomes where the total genome sums up to 30 Mb on each SMRT Cell. As microbial genomes and gDNA samples vary in genetic complexity and quality, respectively, we describe general recommendations and best practices.

Core Lab Brochure: The most trusted long-read technology

2018

Summary +

SMRT Sequencing is Smart Business. Scientists rely on long-read sequencing to generate high-quality reference genomes, more accurate gene and transcript models, and an integrated view of the epigenome for their organisms of interest. Single Molecule, Real-Time (SMRT) Sequencing from PacBio is the most trusted long-read sequencing solution available today, delivering comprehensive genomic information for microbes, complex plant and animal genomes, and human biomedical research.

Technical Note: Preparing libraries for PacBio Whole Genome Sequencing for de novo Assembly – Quality Control and Size Selection

2018

Summary +

Single Molecule, Real-Time (SMRT) Sequencing uses the natural process of DNA replication to sequence long fragments of native DNA. As such, starting with high-quality, high molecular weight genomic DNA will result in longer libraries and better performance across difficult to sequence regions of the genome. This technical note is intended to give recommendations, tips and tricks for assessing and preserving the quality and size of your SMRTbell library, and size selection procedures for libraries intended to be used with whole genome sequencing for de novo assembly.

Technical Note: Preparing DNA for PacBio Whole Genome Sequencing for de novo Assembly – Quality Control and Shearing

2018

Summary +

Single Molecule, Real-Time (SMRT) Sequencing uses the natural process of DNA replication to sequence long fragments of native DNA. As such, starting with high-quality, high molecular weight (HMW) genomic DNA (gDNA) will result in longer libraries and better performance across difficult to sequence regions of the genome. This technical note is intended to give recommendations, tips and tricks for assessing and preserving the quality and size of your gDNA sample, shearing methods, and size selection procedures for samples intended to be used with whole genome sequencing for de novo assembly.

Technical Note: Preparing Samples for PacBio Whole Genome Sequencing for de novo Assembly – collection and storage

2018

Summary +

Single Molecule, Real-Time (SMRT) Sequencing uses the natural process of DNA replication to sequence long fragments of native DNA. As such, starting with high-quality, high molecular weight (HMW) genomic DNA (gDNA) will result in better sequencing performance across difficult to sequence regions of the genome. To obtain the highest quality, long DNA it is important to start with sample types compatible with HMW DNA extraction methods. This technical note is intended to give general guidance on sample collection, preparation, and storage across a range of commonly encountered sample types used for SMRT Sequencing whole genome projects. It is important to note that all samples and projects are unique and may not be comprehensively addressed in this document.

Application Brief: Long-read RNA sequencing – Best Practices.

2018

Summary +

With Single Molecule, Real-Time (SMRT) Sequencing and the Sequel System, you can easily and affordably sequence transcript isoforms of up to 10 kb in their entirety. The Iso-Seq method allows users to generate full-length cDNA sequences - with no assembly required - in order to confidently characterize the full complement of transcript isoforms within targeted genes, or across an entire transcriptome.

Application Brief: Low-coverage, long-read whole genome sequencing for structural variation – Best Practices.

2018

Summary +

With the Sequel System, you can affordably and sensitively characterize structural variation (SV) of all types ranging in size from tens to thousands of base pairs. Low-coverage, long-read whole genome sequencing (WGS) data provides rapid discovery of common SVs for population genetics studies and resolves rare SVs unique to an individual, with a very low false-discovery rate.

Application Brochure: No-Amp targeted sequencing.

2018

Summary +

Due to technology limitations, repeat-expansion disorders have gone without the needed base-level resolution of the disease causative long repetitive elements. Enrichment of these hard-to-amplify genomic regions is now possible with our no-amplification (No-Amp) targeted sequencing method utilizing the CRISPR/Cas9 system.

De Novo Assembly Brochure: Assembly Options for your SMRT Sequencing Data

2018

Summary +

De novo assembly using PacBio SMRT Sequencing is now the gold standard for generating contiguous, highly accurate reference genomes. The challenge of assembly is eased with PacBio long reads, and a wealth of bioinformatics tools have been developed and optimized for SMRT Sequencing. Whether you’re a genome expert who likes to run tools at the command line or you’d prefer assembly experts to deliver a polished assembly directly to you, PacBio has a solution to fit your needs.

Case Study: Pioneering a pan-genome reference collection

2018

Summary +

At DuPont Pioneer, DNA sequencing is paramount for R&D to reveal the genetic basis for traits of interest in commercial crops such as maize, soybean, sorghum, sunflower, alfalfa, canola, wheat, rice, and others. They cannot afford to wait the years it has historically taken for high-quality reference genomes to be produced. Nor can they rely on a single reference to represent the genetic diversity in its germplasm.