Nature biotechnologyOct 5, 2021

Designed pegRNAs increase the accuracy of prime editing

CRISPR Gene Editing Weekly Brief ↗PubMed ↗DOI ↗

Updated Mar 31, 2026

Abstract

Engineered pegRNAs (epegRNAs) improve prime editing efficiency by 3-4 times in various human cell types.

Prime editing allows for precise modifications in DNA, including point mutations and small insertions or deletions.
Degradation of specific regions of the pegRNA can diminish the effectiveness of prime editing.
Incorporating structured RNA motifs at the 3' end of pegRNAs enhances their stability and editing efficiency.
The modified pegRNAs maintain a low risk of off-target effects during DNA editing.
Optimized structural motifs and a new computational tool (pegLIT) were developed to improve pegRNA design.
epegRNAs facilitate the installation or correction of mutations associated with diseases.

Simplified

Prime editing enables the installation of virtually any combination of point mutations, small insertions or small deletions in the DNA of living cells. A prime editing guide RNA (pegRNA) directs the prime editor protein to the targeted locus and also encodes the desired edit. Here we show that degradation of the 3' region of the pegRNA that contains the reverse transcriptase template and the primer binding site can poison the activity of prime editing systems, impeding editing efficiency. We incorporated structured RNA motifs to the 3' terminus of pegRNAs that enhance their stability and prevent degradation of the 3' extension. The resulting engineered pegRNAs (epegRNAs) improve prime editing efficiency 3-4-fold in HeLa, U2OS and K562 cells and in primary human fibroblasts without increasing off-target editing activity. We optimized the choice of 3' structural motif and developed pegLIT, a computational tool to identify non-interfering nucleotide linkers between pegRNAs and 3' motifs. Finally, we showed that epegRNAs enhance the efficiency of the installation or correction of disease-relevant mutations.

Full Text

Full text is available at the source.

View Full Text ↗

Designed pegRNAs increase the accuracy of prime editing

Abstract

Full Text

You found one interesting study. We’ll send the next 7.

what lands in your inbox each week:

Recent issues from the crispr gene editing brief

Abstract

Full Text

Related papers

You found one interesting study. We’ll send the next 7.

what lands in your inbox each week:

Recent issues from the crispr gene editing brief