The challenge of isoform reconstruction
In eukaryotic organisms, the majority of genes are alternatively spliced to produce multiple transcript isoforms, dramatically increasing the protein-coding potential of a genome1. Alternatively spliced isoforms from the same gene can have significantly different, even antagonistic, effects2. To study gene expression, researchers have looked at fragments of an organism’s genes utilizing next-generation sequencing methods, commonly referred to as RNA sequencing (RNA-seq). However, short-read RNA-seq cannot span full-length transcripts, making it difficult to accurately characterize the diverse landscape of isoforms3.
Produce full-length transcripts without assembly
The isoform sequencing (Iso-Seq) application generates full-length cDNA sequences — from the 5’ end of transcripts to the poly-A tail — eliminating the need for transcriptome reconstruction using isoform-inference algorithms. The Iso-Seq method generates accurate information about alternatively spliced exons and transcriptional start sites. It also delivers information about poly-adenylation sites for transcripts up to 10 kb in length across the full complement of isoforms within targeted genes or the entire transcriptome.
PacBio delivers confident characterization of isoform diversity with the PacBio Systems
- Profile the full complexity of the human transcriptome and discover novel genes, isoforms, and gene fusion events
- Survey transcript diversity in plants and animals to improve genome annotation and gene discovery
- Improve quantitation accuracy of short-read RNA-seq data with sample-specific gene isoform models
- Sequence full-length bacterial operons from transcriptomes and metatranscriptomes
To learn more about how to profile the complexity of the transcriptome with the Iso-Seq application, contact us.
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- Kuang, Zheng et al. (2016) The dynamic landscape of fission yeast meiosis alternative-splice isoforms bioRxiv
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- Dong, Lingli et al. (2015) Single-molecule real-time transcript sequencing facilitates common wheat genome annotation and grain transcriptome research. BMC Genomics
- Westbrook, Catherine J et al. (2015) No assembly required: Full-length MHC class I allele discovery by PacBio circular consensus sequencing. Human Immunology
- Lu, Tingting et al. (2015) Transcriptome-wide investigation of circular RNAs in rice. RNA
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- Xu, Zhichao et al. (2015) Full-length transcriptome sequences and splice variants obtained by a combination of sequencing platforms applied to different root tissues of Salvia miltiorrhiza and tanshinone biosynthesis. The Plant Journal
- Tombácz, Dóra et al. (2015) Characterization of novel transcripts in pseudorabies virus. Viruses
- Dasari, Surendra et al. (2015) Proteomic detection of immunoglobulin light chain variable region peptides from amyloidosis patient biopsies. Journal of Proteome Research
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- Cavelier, Lucia et al. (2015) Clonal distribution of BCR-ABL1 mutations and splice isoforms by single-molecule long-read RNA sequencing. BMC Cancer
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- Poster: Tseng, Elizabeth et al. (2016) Reconstruction of the spinach coding genome using full-length transcriptome without a reference genome
- Poster: Baybayan, Primo et al. (2016) Application specific barcoding strategies for SMRT Sequencing
- Poster: Clark, Tyson et al. (2016) Full-length cDNA sequencing for genome annotation and analysis of alternative splicing
- Presentation: Nattestad, Maria et al. (2015) Comprehensive genome and transcriptome structural analysis of a breast cancer cell line using PacBio long read sequencing
- Poster: Clark, T. et al. (2015) Full-length cDNA sequencing of alternatively spliced isoforms provides insight into human cancer
- Poster: Boitano, Matthew et al. (2015) Full-length cDNA sequencing of prokaryotic transcriptome and metatranscriptome samples.
- Poster: Tseng, Elizabeth et al. (2015) Full-length isoform sequencing of the human MCF-7 cell line using PacBio long reads.
- Poster: Clark, Tyson A. et al. (2015) Full-length cDNA sequencing of alternatively spliced isoforms provides insight into human diseases.
- Poster: Baybayan, Primo et al. (2015) SMRT Sequencing of DNA and RNA samples extracted from formalin-fixed and paraffin embedded tissues using adaptive focused acoustics by Covaris.
- Poster: Clark, Tyson A et al. (2015) Single Molecule, Real-Time sequencing of full-length cDNA transcripts uncovers novel alternatively spliced isoforms.
- Poster: Copetti, Dario et al. (2015) Rapid full-length Iso-Seq cDNA sequencing of rice mRNA to facilitate annotation and identify splice-site variation.
- Timp, Winston (2016) PAG Conference: An extreme metabolism: Iso-Seq analysis of the ruby-throated hummingbird transcriptome
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- Clark, Tyson (2015) Seminar: Iso-seq method – sample prep and experimental design for full-length cDNA sequencing
- Smith, Tim (2015) Customer Experience: Long-read assemblies correct problems, advance research in livestock
- Racacho, Lemuel (2015) Customer Experience: For long, noncoding RNAs, SMRT Sequencing offers better view
- Tseng, Elizabeth (2015) Seminar: Iso-seq analysis & beyond – advanced bioinformatics for transcriptome sequencing using long reads
- Nattestad, Maria (2015) ASHG Conference: Comprehensive genome and transcriptome structural analysis of a breast cancer cell line using PacBio long read sequencing
- Gyllensten, Ulf (2015) AGBT Virtual Poster: Clinical sequencing using Pacific Biosciences RS II for HLA typing and monitoring of drug resistance in chronic myeloid leukemia (CML)
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