A long-read RNA-seq approach to identify novel transcripts of very large genes

July 2020

Prech Uapinyoying, Jeremy Goecks, Susan M. Knoblach, Karuna Panchapakesan, Carsten G. Bonnemann, Terence A. Partridge, Jyoti K. Jaiswal, and Eric P. Hoffman

Scientists researching muscular disorders present a method for studying gene expression of large RNA transcripts. The PacBio Iso-Seq method allows the sequencing of large transcripts for the identification of isoforms and alternative splicing. However, it has not been amenable to accurate quantitation of expression. Here the scientists combine Iso-Seq with PacBio’s HiFi high consensus accuracy method to show differential gene expression of large transcripts in muscles as well as identify and quantitate previously unannotated ones.

BluePippin was used to size select 5-10kb cDNA fragments for PacBio Iso-Seq library construction, and a second size selection step to remove unligated adapters.

Author Affiliations:
Center for Genetic Medicine Research, Children’s Research Institute, Washington D.C.

Department of Genomics and Precision Medicine, The George Washington University School of Medicine and Health Sciences, Washington, D.C.
National Institute of Neurological Disorders and Stroke, NIH, Bethesda, MD

Computational Biology Program, Oregon Health and Science University, Portland, OR

Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, Binghamton University, Binghamton, NY

Genome Research
DOI: 10.1101/gr.259903.119.

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