bioRxiv : the preprint server for biologyMar 23, 2026

Spligation allows controlled creation of mixed RNA molecules inside living cells

CRISPR Gene Editing Weekly Brief ↗PubMed ↗DOI ↗OA ↗

Updated Jun 27, 2026

Abstract

Essence

Spligation used CRISPR-Csm cleavage and RNA ligation to make programmable transcript deletions and chimeric RNAs inside living cells.

Evidence

Cell-based RNA engineering experiments tested type III-A CRISPR-Csm complexes, Csm-RtcB fusions, two-transcript cleavage, and endogenous transcript recombination in human cells.

Caveat

The abstract demonstrates molecular feasibility in cells but does not quantify therapeutic performance, specificity, delivery, or disease outcomes.

Simplified

The ability to precisely modify RNA offers opportunities to manipulate the flow of genetic information and influence transcript stability, localization and translation. RNA-targeting technologies enable RNA knockdown, base editing and-splicing, but more extensive transcript changes typically require genome editing or rely on the endogenous splicing machinery. Based on the ability of type III-A CRISPR-Csm complexes to catalyze programmable RNA cleavage in human cells, we investigated their potential to induce site-specific deletions while leaving the remainder of the transcript intact. Our data show that CRISPR-Csm complexes can generate short and long RNA excisions within a target transcript, and that the efficiency of this process is enhanced by fusion of Csm to the RNA ligase RtcB. Furthermore, cleavage of two different transcripts can trigger subsequent-ligation of the cleaved products into a chimeric transcript ("spligation"). Finally, we apply spligation to endogenous transcripts, using Csm to generate recombinant mRNA in cells independent of canonical splice sites. Collectively, this approach enables new forms of precise RNA manipulation in cells with potential applications in human disease. trans trans