# A Unified Nanopore Platform for Direct Sequencing of Individual Full Length RNA Strands Bearing Modified Nucleotides

> **NIH NIH R01** · UNIVERSITY OF CALIFORNIA SANTA CRUZ · 2021 · $954,816

## Abstract

ABSTRACT
We propose to implement a unified platform for detecting RNA nucleotide modifications on contiguous full
length RNA strands using nanopore direct RNA sequencing. In principle, this technology could be used for all
classes of RNA molecules in prokaryotic and eukaryotic cells. It is a logical technical advance for RNA
sequencing because the current standard, sequencing-by-synthesis using Solexa technology requires
conversion of native cellular RNA into short (~300 nt) cDNA amplicons. In so doing, nucleotide modifications
are erased and the continuity of intact RNA strands is lost, thus precluding accurate quantification of RNA
isoforms. Nanopore strand sequencing overcomes these limitations because the ~2 nanometer-long integral
sensor touches and identifies each base along intact native RNA strands as they are driven through the pore
by an applied voltage. Thus, end-to-end sequence analysis of a given RNA strand is achieved.
During the grant period, we will pursue three specific aims: 1) Establish baseline performance of ONT direct
RNA sequencing, and implement targeted improvements; 2) Implement methods to discover and document
nucleotide modifications on native RNA strands; and 3) Optimize nanopore technology for analysis of mRNA
isoform diversity.
UC Santa Cruz is uniquely equipped to undertake this project:
i) We pioneered nanopore RNA strand analysis (Akeson/Deamer) and recently demonstrated that the Oxford
Nanopore MinION nanopore sequencer can resolve single nucleotide variants and base modifications in single
16S rRNA strands (Akeson).
ii) Our RNA Center includes fourteen faculty members. Co-investigators on this application have expertise in
mechanisms of RNA splicing (Sanford), the functional consequences of normal and aberrant RNA isoform
synthesis (Brooks), and the structural biology of RNA (Ares). Collaborating RNA Center faculty will advise on
nuclear non-coding RNA experiments (Carpenter, Kim) and on H9 ES cells (Salama).
iii) Our Genomics Institute is internationally recognized for bioinformatics, including recent advances in
application of Recurrent Neural Networks to human genome haplotyping, and genome-wide nanopore
detection of base modifications (Paten).

## Key facts

- **NIH application ID:** 10163247
- **Project number:** 5R01HG010053-04
- **Recipient organization:** UNIVERSITY OF CALIFORNIA SANTA CRUZ
- **Principal Investigator:** MARK A AKESON
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $954,816
- **Award type:** 5
- **Project period:** 2018-06-01 → 2024-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10163247

## Citation

> US National Institutes of Health, RePORTER application 10163247, A Unified Nanopore Platform for Direct Sequencing of Individual Full Length RNA Strands Bearing Modified Nucleotides (5R01HG010053-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10163247. Licensed CC0.

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