Nanopore native RNA sequencing of a human poly(A) transcriptome

By R. Workman, Alison D. Tang, Paul S. Tang, Miten Jain, John R. Tyson, Roham Razaghi, P. Zuzarte, Timothy Gilpatrick, Alexander Payne, Joshua Quick and 14 more. Nature Methods, 2018

631 citations recorded by the source corpus.

High-throughput complementary DNA sequencing technologies have advanced our understanding of transcriptome complexity and regulation. However, these methods lose information contained in biological RNA because the copied reads are often short and modifications are not retained. We address these limitations using a native poly(A) RNA sequencing strategy developed by Oxford Nanopore Technologies. Our study generated 9.9 million aligned sequence reads for the human cell line GM12878, using thirty MinION flow cells at six institutions. These native RNA reads had a median length of 771 bases, and a maximum aligned length of over 21,000 bases. Mitochondrial poly(A) reads provided an internal measure of read-length quality. We combined these long nanopore reads with higher accuracy short-reads and annotated GM12878 promoter regions to identify 33,984 plausible RNA isoforms. We describe strategies for assessing 3′ poly(A) tail length, base modifications and transcript haplotypes. Directly sequencing RNA strands through a nanopore retains the full length of the transcript and allows for analysis of polyA tail length, transcript haplotypes and base modifications.

Subfields: Molecular Biology

View Paper: https://www.semanticscholar.org/paper/183f9fcc229ce4adf8ac76332092ea977bc4fe65

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Citations in the corpus

• Long-read human genome sequencing and its applications (2020, neutral): “Full-length Full-length poly(A) transcriptomes have been readily obtained by ONT native RNA sequencing 123,124 .”
• Nanopore sequencing technology, bioinformatics and applications (2021, background): “Since then, epitranscriptome analyses using ONT sequencing have progressed rapidly, including detection of 7-methylguanosine (m 7 G) and pseudouridine in 16S rRNAs of E. coli 183 , m 6 A in mRNAs of S. cerevisiae 73 and A. thaliana 168 and m 6 A 130 and pseudouridine 104 in human RNAs.”