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Obtain a more comprehensive set of transcript isoforms

A single gene may code for a surprising number of proteins, each with a distinct biological function. This is especially true in higher organisms. Short-read RNA sequencing (RNA-seq) works by breaking up transcript isoforms into smaller pieces and bioinformatically reassembling them, leaving opportunity for misassembly or incomplete capture of the full diversity of isoforms from genes of interest.

Capture full-length transcripts with no assembly required

The PacBio Isoform Sequencing (Iso-Seq) method employs long reads to sequence transcript isoforms of up to 10 kb in their entirety. Surveying this transcript diversity, either broadly or in a targeted fashion, reveals key information about the frequency and types of alternative transcription, improving genome annotation and gene discovery.

Single Molecule, Real-Time (SMRT) Sequencing enhances RNA sequencing, allowing you to:

  • Make direct, evidence-based genome annotations
  • Discover novel genes and isoforms, even in well-characterized samples
  • Identify promoters and splice sites in comprehensive gene models to understand gene regulation
  • Improve accuracy of RNA-seq quantification with isoform-level resolution for gene expression studies
  • Distinguish important stress response, developmental, and tissue-specific isoforms

Workflow: from RNA to full-length transcripts

  • SMRT Sequencing with PacBio Systems
    • Simplify sample prep workflow and reduce project costs by using the Sequel System
    • Directly sequence full-length transcripts with average read lengths of ~10 kb
    • Scale throughput based on project needs
    • Scalable throughput
      • Profile multiplexed transcripts in a single SMRT Cell
      • Survey transcriptomes in 1–2 SMRT Cells on the Sequel System
      • Increase sequencing depth for more comprehensive transcriptome characterization

Data Release

 

Iso-Seq Results for Hummingbird and Zebra Finch Brain Tissue

Explore the role of the transcriptome in avian vocal learning with the PacBio Iso-Seq data set generated with the Sequel System.

To learn more and access the data, see our blog post.

Iso-Seq Project Calculator

 

Tell us about your plant or animal RNA sequencing project and receive an estimate on the time and materials needed using the Iso-Seq method on the Sequel System.


Spotlight: Significantly improved sorghum genome annotation

Sorghum genome annotation

Splice isoform analysis in the sorghum transcriptome using the Iso-Seq method greatly improved genome annotation, with >77,000 novel splice isoforms and >2,100 novel genes identified. In this example, a gene was discovered to produce 13 novel alternatively spliced isoforms, where the previous gene model contained only a single isoform. Explore this research further:

Abdel-Ghany, S.E. et al. (2016) A survey of the sorghum transcriptome using single-molecule long reads. Nature Communications. 7, e11706.

To learn more about how to profile the complexity of the transcriptome with the Iso-Seq method, contact us.

Selected Resources