New Solutions for Comprehensive and Efficient Targeted Sequencing and Multiplexing of Samples
Monday, April 27, 2015
We are proud to announce the introduction of several new solutions for targeted sequencing and sample multiplexing on the PacBio® Sequencing System.
New Targeted Sequencing Workflow through Collaboration with Roche NimbleGen
Today we announced a new workflow that combines Roche NimbleGen’s SeqCap EZ enrichment technology with large DNA fragments (up to 6 kb) and our Single Molecule, Real-Time (SMRT®) Sequencing to provide a more comprehensive view of variants, transgene integration sites, and haplotype information over multi-kilobase contiguous regions. The laboratory workflow is described in a shared protocol. For each targeted region, SAMtools are used to phase and bin reads by haplotype, and then Quiver is applied to polish each haplotype to high consensus accuracy. This entire bioinformatics workflow is summarized on GitHub.
The long fragments capture haplotype information that provides a more inclusive view of the targeted region of interest, allowing a comprehensive look at variants in exons, introns, and intergenic regions, as well as the generation of accurate haplotypes as demonstrated in this application note. In a recent publication by Richard Gibbs and colleagues from Baylor College of Medicine, this new enrichment method was used for de novo sequencing and detection of structural variation involved in human genetic disease.
“The use of NimbleGen Sequence Capture technology to enrich large DNA molecules for targeted sequencing on the PacBio platform leverages the advanced capabilities of both systems to achieve unprecedented efficiency for targeted genetic phasing and structural variant elucidation,” said Rebecca Selzer, Ph.D., President, Roche NimbleGen. “The ability to focus SMRT Sequencing onto discrete regions of interest within a genome demonstrates that the technical advantages inherent in long, single molecule sequencing can be part of a higher-throughput analysis strategy.”
Multiplexing and Barcoding Workflows
We have also introduced two multiplexing workflows and associated products for barcoding samples prior to running on SMRT Cells. The new barcoding kits allow you to pool many samples and thus reduce the cost per sample and the total time for sample preparation. By employing the barcoding strategies, you can efficiently focus on even more defined genomic regions. And by barcoding samples, you have the flexibility to multiplex samples or targets within a sample, or a combination of both.
Kevin Corcoran, Senior Vice President of Market Development for Pacific Biosciences, commented: “The new barcoding methods allow a greater numbers of samples to be studied simultaneously and also improve the efficiency of SMRT Sequencing, by increasing SMRT Cell sample capacity and streamlining the workflow.”