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Single-molecule, full-length transcript isoform sequencing reveals disease-associated RNA isoforms in cardiomyocytes.

Department of Genetics, School of Medicine, Stanford University, Stanford, USA. European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany. Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, USA. Collaboration for joint PhD degree between EMBL and Heidelberg University, Faculty of Biosciences, Heidelberg, Germany. Institute for Cardiomyopathies Heidelberg (ICH), Heart Center Heidelberg, University of Heidelberg, Heidelberg, Germany. DZHK (German Center for Cardiovascular Research), partner site Heidelberg, Heidelberg, Germany. Department of Medicine III, University of Heidelberg, Heidelberg, Germany. CAS Key Laboratory of Computational Biology, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai, China. wuwei@picb.ac.cn. Center for Biomedical Informatics, Shanghai Engineering Research Center for Big Data in Pediatric Precision Medicine, Shanghai Children's Hospital, Shanghai Jiao Tong University, Shanghai, China. wuwei@picb.ac.cn. Stanford Genome Technology Center, Stanford University, Palo Alto, USA. wuwei@picb.ac.cn. Department of Genetics, School of Medicine, Stanford University, Stanford, USA. larsms@stanford.edu. European Molecular Biology Laboratory (EMBL), Genome Biology Unit, Heidelberg, Germany. larsms@stanford.edu. Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, USA. larsms@stanford.edu. Stanford Genome Technology Center, Stanford University, Palo Alto, USA. larsms@stanford.edu. DZHK (German Center for Cardiovascular Research), partner site EMBL Heidelberg, Heidelberg, Germany. larsms@stanford.edu.

Nature communications. 2021;(1):4203
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Abstract

Alternative splicing generates differing RNA isoforms that govern phenotypic complexity of eukaryotes. Its malfunction underlies many diseases, including cancer and cardiovascular diseases. Comparative analysis of RNA isoforms at the genome-wide scale has been difficult. Here, we establish an experimental and computational pipeline that performs de novo transcript annotation and accurately quantifies transcript isoforms from cDNA sequences with a full-length isoform detection accuracy of 97.6%. We generate a searchable, quantitative human transcriptome annotation with 31,025 known and 5,740 novel transcript isoforms ( http://steinmetzlab.embl.de/iBrowser/ ). By analyzing the isoforms in the presence of RNA Binding Motif Protein 20 (RBM20) mutations associated with aggressive dilated cardiomyopathy (DCM), we identify 121 differentially expressed transcript isoforms in 107 cardiac genes. Our approach enables quantitative dissection of complex transcript architecture instead of mere identification of inclusion or exclusion of individual exons, as exemplified by the discovery of IMMT isoforms mis-spliced by RBM20 mutations. Thereby we achieve a path to direct differential expression testing independent of an existing annotation of transcript isoforms, providing more immediate biological interpretation and higher resolution transcriptome comparisons.