# Universally-applicable RNA mapping at subcellular and single-base resolution

> **NIH NIH DP2** · UNIVERSITY OF CALIFORNIA-IRVINE · 2022 · $1,354,500

## Abstract

Project Summary/Abstract
Each step-change in increasing resolution on RNA identification and quantification (i.e., transcriptome profiling)
has been transformative to our understanding of cell biology. As the most recent technological revolution, the
ability to map the spatial distribution of RNA molecules has revealed the importance of subcellular RNA
localization, advanced our understanding of cell and developmental biology, and transformed the single cell
transcriptomics field. However, despite the recent development, current RNA-mapping tools are limited to
molecules with long specific sequence (mostly > 500 nt). Many other important RNA species (such as splicing
isoforms, miRNA, and RNA editing), with much shorter specific sequence motif, remain inaccessible in situ. Our
goal is to make this next resolution jump in profiling transcriptomics, i.e., RNA-mapping with high transcript-
selectivity and single base-sensitivity. We plan to achieve the goal by developing a new tool, named SMOOTHY-
FISH (Single-nucleotide specificity, Minimum Of Off-Target effects, and High Yield), which enables spatially
mapping all types of RNA molecules in individual cells. In this proposal, we will demonstrate SMOOTHY-FISH
from two aspects: technique development and one example application test. More specifically, for the technical
development, we anticipate achieving a platform that can suggest robust and reliable SMOOTHY-FISH probe-
sets to any assigned RNA target sequence, a user-friendly system that will be widely accessible. To achieve the
goal, we will incorporate the unique power of magnetic tweezers (a single molecule tool with nanometer and
milliseconds resolution) to systemically quantify the dynamics and kinetics of nucleic acid hybridization under
various conditions, to reveal how fast and how stable the designed SMOOTHY-FISH probe-sets can form the
needed secondary structure, and to build a mechanistic modeling framework to describe and predict the
hybridization dynamic process. As for the example application test, we will use SMOOTHY-FISH to study the
mystery biogenesis process of circular RNA with isoform-specificity (i.e., co-existence with its linear isoforms)
for the first time. We anticipate revealing the potential to modulate circular RNA expression quantitatively and
efficiently, i.e., lay a foundation for therapeutic strategies in the future. Together, this work will provide a next-
generation tool for in situ RNA identification and quantification, allow us to localize and quantify many important,
but currently inaccessible RNA species at the single molecule level in individual cells, thus help open a new era
of RNA biology, from basic science research to clinical applications.

## Key facts

- **NIH application ID:** 10473389
- **Project number:** 1DP2GM149554-01
- **Recipient organization:** UNIVERSITY OF CALIFORNIA-IRVINE
- **Principal Investigator:** Fangyuan Ding
- **Activity code:** DP2 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $1,354,500
- **Award type:** 1
- **Project period:** 2022-09-01 → 2025-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10473389

## Citation

> US National Institutes of Health, RePORTER application 10473389, Universally-applicable RNA mapping at subcellular and single-base resolution (1DP2GM149554-01). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10473389. Licensed CC0.

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