# Molecular basis for CRISPR RNA-guided nucleic acid cleavage and DNA transposition

> **NIH NIH R01** · PURDUE UNIVERSITY · 2022 · $331,700

## Abstract

PROJECT SUMMARY
CRISPR-Cas systems provide adaptive immunity in bacteria and archaea by employing guide RNAs and
endonuclease effectors to specifically recognize and cleave invasive nucleic acids. The specific DNA targeting
and cleavage activities of CRISPR-Cas systems have been adopted and developed for genome editing and
various other applications, which are revolutionizing biomedical research and beyond. However, safety
concerns are raised because of off-target genome editing and the dependence of these systems on
endogenous host DNA repair pathways, hindering clinical application. Exploration of alternative CRISPR-Cas
systems in nature not only offers an opportunity to overcome those challenges but may also inspire new
applications. Structural and biochemical characterizations of CRISPR-Cas systems are critical for
understanding their mechanisms and repurposing them for precise genome editing. Our long-term goals are to
unravel the mechanisms underlying target nucleic acid recognition and cleavage mediated by type V and
transposon-associated CRISPR-Cas systems, which provide essential knowledge for safer and more reliable
application in treating human disease. In this proposal, we will work on the molecular mechanisms for four
newly discovered CRISRP-Cas systems, covering DNA targeting (Cas12i), RNA targeting (Cas12g), and
CRISPR RNA-guided DNA transposition (type I-F Cascade and Cas12k). As revealed in our preliminary data,
Cas12i accommodates a longer crRNA-DNA heteroduplex than currently used Cas effectors, thus potentially
improving specificity for genome editing. The RNA-guided RNase Cas12g is compact and thermostable,
highlighting its potential for RNA editing and RNA targeting. Furthermore, type I-F Cascade and Cas12k direct
transposition machinery for RNA-guided DNA transposition, opening a new paradigm for genome editing
independent of DNA repair pathways.
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## Key facts

- **NIH application ID:** 10455694
- **Project number:** 5R01GM138675-03
- **Recipient organization:** PURDUE UNIVERSITY
- **Principal Investigator:** Leifu Chang
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $331,700
- **Award type:** 5
- **Project period:** 2020-08-01 → 2025-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10455694

## Citation

> US National Institutes of Health, RePORTER application 10455694, Molecular basis for CRISPR RNA-guided nucleic acid cleavage and DNA transposition (5R01GM138675-03). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10455694. Licensed CC0.

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