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bioRxiv · 10.1101/2022.04.05.487236

Enhancing Prime Editing Efficiency and Flexibility with Tethered and Split pegRNAs

Feng, Y.·Wang, S.·Mo, Q.·Xiao, X.·Liu, P.·Ma, H.
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Abstract

Prime editing (PE) has advantages for small insertion, deletion or point mutations without double-stranded DNA breaks. The 3-extension of pegRNAs could negatively affect its stability or folding and comprise the PE activity. Here we generated stem-loop PEs (sPEs) by adding stem-loop aptamers at the 3-terminal of pegRNA, which can be tethered to Cas9 nickase resulting in tethered PEs (tPEs). sPEs and tPEs increased the small insertion, deletion or point mutations efficiency by 2-4-fold on average in HEK293, U2OS and HeLa cells. We split the modified pegRNAs into sgRNA and prime RNA. The resulting split pegRNA prime editors (SnPEs) maintain the PE activity and increase flexibility.

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Feng, Y., Wang, S., Mo, Q., Xiao, X., Liu, P., Ma, H.. 2022-04-05. Enhancing Prime Editing Efficiency and Flexibility with Tethered and Split pegRNAs. https://doi.org/10.1101/2022.04.05.487236

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